Anti-Microbial Immunomodulation

ABSTRACT

The invention provides methods of modulating an immune system in a vertebrate host for the therapeutic or prophylactic treatment of infection by a first microbial pathogen in a target tissue, comprising administration at an administration site of an effective amount of an antigenic formulation comprising antigenic determinants specific for a second heterologous microbial pathogen.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/308,302 filed Nov. 1, 2016, which application is a U.S. NationalPhase Application of PCT Application No. PCT/CA2015/050377 filed May 1,2015, which application claims priority to U.S. Provisional PatentApplication Ser. No. 61/988,117 filed May 2, 2014, the disclosure ofwhich are herein incorporated by reference.

FIELD OF THE INVENTION

In various aspects, the invention relates to immunological therapies fortreating or preventing pathologies associated with microbial infectionsin a vertebrate, including the use of microbial vaccines.

BACKGROUND OF THE INVENTION

The innate immune system and the adaptive immune system work in concertin vertebrates to provide, among many other things, protection frompathogenic infection by micro-organisms. Anti-microbial vaccines may beformulated to engage both the innate and adaptive immune systems, but aneffective response to vaccination is generally understood to involve aspecific adaptive response to one or more of the immunogens present in avaccine. In this way, multivalent vaccines, such as some pneumococcalvaccines, may be used to elicit a specific adaptive response to morethan one serovar. Vaccines have also been described that confer somedegree of cross-protective immunity, in which cross-reactivity to anantigen other than the immunogen confers a degree of protective immunityto heterologous microorganisms.

SUMMARY OF THE INVENTION

In one aspect, the invention provides methods and compositions fortreating a vertebrate subject for a condition characterized bypathologies associated with a microbial infection, involving the use ofmicrobial vaccines derived from one pathogenic organism to treatinfections caused by a heterologous pathogenic organism.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A, 1B and 1C show the number of inflammatory monocytes anddendritic cells in the draining lymph node, lungs, and spleen of micefollowing treatment with either a K. pneumoniae antigenic composition orPBS, as described in Example 1A herein.

FIG. 2 shows the total number of monocytes and dendritic cells in thelung, peritoneum and spleen of mice following treatment with either a K.pneumoniae antigenic composition, an E. coli antigenic composition, orPBS, as described in Example 1B herein.

FIG. 3 shows the total number of CD4+ T cells, CD8+ T cells, and NKcells from mice treated with either a K. pneumoniae antigeniccomposition, an E. coli antigenic composition, or PBS, as described inExample 1B herein.

FIGS. 4A and 4B show the total number of (A) inflammatory monocytes anddendritic cells and (B) CD4+ T cells, CD8+ T cells, and NK cells frommice treated with either a heat-inactivated K. pneumoniae antigeniccomposition, a phenol-inactivated K. pneumoniae antigenic composition,or PBS, as described in Example 10 herein.

FIG. 5 shows the relative frequency of CD11b+ Gr-1+ cells detected fromthe colons of mice treated with either K. pneumoniae or E. coliantigenic compositions or with PBS control.

FIGS. 6A and 6B show the relative frequency of CD11 b+ Gr-1+ cellsdetected from the lungs of mice treated with either K. pneumoniae or E.coli antigenic compositions or with PBS control.

FIGS. 7A to 7E illustrate microbial prophylaxis as discussed in Example3, in which treatment with a composition comprising whole killed K.pneumoniae cells provides protective immunity against subsequentchallenge by S. pneumoniae. FIG. 7A is a survival curve 5 dayspost-challenge. FIGS. 7B-7E illustrate S. pneumoniae quantified in nasalwash, lung and spleen homogenate. Megakaryocytes were counted in tenadjacent high power fields (400× magnification) of histological spleensections. Statistical significance was determined by two-tailedMann-Whitney test.

FIGS. 8A and 8B illustrate heterologous anti-microbial therapy in amouse model, as discussed in Example 4, in which treatment with acomposition comprising whole killed E. coli cells is effective inameliorating an infection caused by a heterologous strain of adherentinvasive E. coli.

FIGS. 9A and 9B illustrate heterologous anti-microbial therapy in amouse model, as discussed in Example 3, in which treatment with acomposition comprising whole killed Klebsiella pneumonia cells iseffective in ameliorating a lung infection caused by Pseudomonasaeruginosa (PA14).

FIG. 10 illustrates heterologous anti-microbial therapy in a mousemodel, as discussed in Example 3, in which treatment with a compositioncomprising whole killed Klebsiella pneumonia cells is effective inameliorating a lung infection caused by Streptococcus pneumoniae(P1542).

FIGS. 11A and 11B illustrate heterologous anti-microbial therapy in amouse model, as discussed in Example 5, in which treatment with acomposition comprising whole killed E. coli cells is effective inameliorating a peritoneal infection caused by S. enterica.

FIGS. 12A, 12B and 12C illustrate heterologous anti-microbial therapy ina mouse model, as discussed in Example 5, in which the effectiveness oftreatments with two different compositions comprising alternativestrains of whole killed E. coli cells is compared to a treatment with anantigenic S. enterica composition in ameliorating a peritoneal infectioncaused by S. enterica.

FIG. 13 is a bar graph illustrating heterologous anti-microbial therapyin a mouse model, as discussed in Example 6, in which the effectivenessof a S. aureus-derived SSI (QBSAU) is shown against P. aeruginosachallenge in skin, with significantly reduced P. aeruginosa bacterialcounts following QBSAU pretreatment.

FIG. 14 is a bar graph illustrating heterologous anti-microbial therapyin a mouse model, as discussed in Example 6, repeating and confirmingthe data shown in FIG. 13, in which the effectiveness of a S.aureus-derived SSI (QBSAU) is shown against P. aeruginosa challenge inskin, with significantly reduced P. aeruginosa bacterial countsfollowing QBSAU pretreatment.

FIG. 15 is a bar graph graph illustrating heterologous anti-microbialtherapy in a mouse model, as discussed in Example 3c, illustratingKlebsiella pneumoniae SSI (QBKPN) demonstrated statistically superiorefficacy in prophylaxis compared to the E. coli SSI (QBECO), inprotecting against P. aeruginosa challenge in the lungs.

FIG. 16 is is a bar graph graph illustrating heterologous anti-microbialtherapy in a mouse model, as discussed in Example Example 3c,illustrating Klebsiella pneumoniae SSI (QBKPN) demonstratedstatistically superior efficacy in prophylaxis compared to the E. coliSSI (QBECO), in protecting against S. pneumoniae challenge in the lungs.

FIGS. 17A and 17B are two graphs showing targeted heterologousanti-microbial therapy in a geriatric mouse model of lung infection,showing that Klebsiella pneumoniae SSI (QBKPN) protects against S.pneumoniae challenge in lungs of aged mice. Survival benefit for oldmice, FIG. 17B, was more pronounced than for young mice, FIG. 17A.

FIG. 18 is a graph illustrating that QBKPN SSI reduces weight loss inaged mice, following challenge with S. pneumoniae, and this benefit isgreater for aged mice compared to young mice.

FIG. 19 illustrates antimicrobial prophylaxis in the peritoneal cavity,with a graph illustrating that both QBECO and QBKPN SSIs are protective,as measured by bacterial load in the spleen, with counts in mice treatedwith QBECO being significantly lower than mice treated with QBKPN (dataare CFU/ml (mean+/−std dev).

DETAILED DESCRIPTION OF THE INVENTION

In various aspects, the invention relates to the surprising discoverythat administration of formulations that include antigenic determinantsof microbial pathogens that are pathogenic in a particular tissue ororgan, is effective in treating pathologies associated with heterologousmicrobial infections in that specific tissue or organ. Compositions ofthe invention may for example be administered at a site that is distantfrom the site of infection. Accordingly, the invention providesantigenic compositions derived from these microbial pathogens, includingwhole killed bacterial, viral or fungal species, or components thereof,for the therapeutic or prophylactic treatment of heterologous microbialinfections, and methods for using the same. The compositions may forexample be derived from endogenous pathogens or exogenous pathogens, asdescribed in more detail below.

Antigenic compositions of the invention may be produced that includeantigenic determinants that together are specific for or characteristicof a microbial pathogen. In this context, by “specific”, it is meantthat the antigenic determinants are sufficiently characteristic of thepathogen that they could be used to raise an immune response, such as anadaptive immune response, against the pathogen in the patient, if theantigenic determinants were to be administered in an appropriate mannerto have that effect. It will be recognized that the antigenicdeterminants need not be so specific that they are characteristic ofonly one particular strain or species of pathogen, since even a specificimmune response against a particular pathogen may be cross reactive withother closely related organisms that are also naturally pathogenic inthe tissue or organ in which the heterologous infection is situated andthat the antigenic composition is formulated or selected to target.

A “cell” is the basic structural and functional unit of a livingorganism. In higher organisms, e.g., animals, cells having similarstructure and function generally aggregate into “tissues” that performparticular functions. Thus, a tissue includes a collection of similarcells and surrounding intercellular substances, e.g., epithelial tissue,connective tissue, muscle, nerve. An “organ” is a fully differentiatedstructural and functional unit in a higher organism that may be composedof different types of tissues and is specialized for some particularfunction, e.g., kidney, heart, brain, liver, etc. Accordingly, by“specific organ, tissue, or cell” is meant herein to include anyparticular organ, and to include the cells and tissues found in thatorgan.

“Pathogenic” agents are agents, such as microbes, such as bacteria orviruses, which are known to cause infection in a host in nature, and inthis sense, “pathogenic” is used in the context of the present inventionto mean “naturally pathogenic”. Although a wide variety of microbes maybe capable of causing infection under artificial conditions, such asartificial inoculations of a microbe into a tissue, the range ofmicrobes that naturally cause infection is necessarily limited, and wellestablished by medical practice.

An “infection” is the state or condition in which the body or a part ofit is invaded by a pathogenic agent (e.g., a microbe, such as abacterium) which, under favorable conditions, multiplies and produceseffects that are injurious (Taber's Cyclopedic Medical Dictionary, 14thEd., C. L. Thomas, Ed., F. A. Davis Company, PA, USA). An infection maynot always be apparent clinically and may result in only localizedcellular injury. Infections may remain subclinical, and temporary if thebody's defensive mechanisms are effective. Infections may spread locallyto become clinically apparent as an acute, a subacute, or a chronicclinical infection or disease state. A local infection may also becomesystemic when the pathogenic agent gains access to the lymphatic orvascular system. Infection is usually accompanied by inflammation, butinflammation may occur without infection.

“Inflammation” is the characteristic tissue reaction to injury (markedby swelling, redness, heat, and pain), and includes the successivechanges that occur in living tissue when it is injured. Infection andinflammation are different conditions, although one may arise from theother (Taber's Cyclopedic Medical Dictionary, supra). Accordingly,inflammation may occur without infection and infection may occur withoutinflammation (although inflammation typically results from infection bypathogenic bacteria or viruses). Inflammation is characterized by thefollowing symptoms: redness (rubor), heat (calor), swelling (infection),pain (dolor). Localized visible inflammation on the skin may be apparentfrom a combination of these symptoms, particularly redness at a site ofadministration.

Various subjects may be treated in accordance with alternative aspectsof the invention. As used herein, a “subject” is an animal, for e.g, avertebrate such as a mammal, to whom the specific pathogenic bacteria,bacterial antigens, viruses, viral antigens or compositions thereof ofthe invention may be administered. Accordingly, a subject may be apatient, e.g., a human, suffering from microbial infection, or suspectedof having a microbial infection, or at risk for developing a microbialinfection. A subject may also be an experimental animal, e.g., an animalmodel of infection. In some embodiments, the terms “subject” and“patient” may be used interchangeably, and may include a human, anon-human mammal, a non-human primate, a rat, mouse, dog, etc. A healthysubject may be a human who is not suffering from an infection orsuspected of having an infection, or who is not suffering from a chronicdisorder or condition. A “healthy subject” may also be a subject who isnot immunocompromised. By “immunocompromised” or “immunosuppressed” ismeant any condition in which the immune system functions in an abnormalor incomplete manner, for example wherein the host is a patient who doesnot have the ability to respond normally to an infection due to animpaired or weakened immune system.

Immunocompromisation or immunosuppression may be due to disease, certainmedications (such as chemotherapeutics used in cancer treatment), orconditions present at birth. Immunocompromised subjects may be foundmore frequently among infants, the elderly, and individuals undergoingextensive drug or radiation therapy. Accordingly, aspects of theinvention involve the treatment of pediatric and geriatric patients, orpatients at risk of a nosocomial infection. Particular patientpopulations may for example include patients with compromised immunesystems due to HIV infection or AIDS, cancer, solid organtransplantation, stem cell transplantation, sickle cell disease orasplenia, congenital immune deficiencies, chronic inflammatoryconditions, cochlear implants, or cerebrospinal fluid leaks.

An “immune response” includes, but is not limited to, one or more of thefollowing responses in a mammal: induction or activation of antibodies,neutrophils, monocytes, macrophages (including both M1-like macrophagesand M2-like macrophages as described herein), B cells, T cells(including helper T cells, natural killer cells, cytotoxic T cells, γδ Tcells), such as induction or activation by the antigen(s) in anantigenic composition, following administration of the composition. Animmune response to a composition thus generally includes the developmentin the host animal of a cellular and/or antibody-mediated response tothe composition of interest. In some embodiments, the immune response issuch that it will also result in slowing or stopping the progression ofan infection in the animal. An immune response includes both cellularimmune responses and humoral immune responses, of both the innate andadaptive immune systems.

In selected embodiments, the methods of the invention may involvedetermining whether the individual has previously been infected with apathogen that is pathogenic in the specific organ or tissue; andadministering to the individual a therapeutic composition comprisingantigenic determinants that are selected or formulated so that togetherthey are specific for the at least one pathogen.

In another aspect, the invention provides methods of formulatingcompositions of the invention for treating an individual for a conditioncharacterized by microbial disease or infection in a specific organ ortissue. The methods may involve determining whether the individual haspreviously been infected with at least one pathogen that is pathogenicin the specific organ or tissue; producing an antigenic compositioncomprising antigenic determinants that together are specific for the atleast one pathogen; and formulating the antigenic composition foradministration as a therapeutic or anti-microbial composition capable ofeliciting an immunological or anti-microbial response in the specificorgan or tissue to a heterologous micro-organism.

The methods detailed herein to determining whether a subject haspreviously been exposed to a pathogen may involve identifying thepresence of at least one antibody that recognizes the pathogen. Themethods may also or alternatively involve identifying at least onememory B cell that recognizes the pathogen. The methods may also oralternatively involve identifying at least one memory T cell thatrecognizes the pathogen. The methods may for example involve obtainingthe antibody, the memory B cell, or the memory T cell from peripheralcirculation or from the targeted specific organ or tissue of theindividual.

In another aspect, the invention provides methods of prophylacticallytreating an individual for an infection in a specific organ or tissue,involving the administration of an infectious micro-organism to provokea potentiating infection in that organ or tissue. The method may forexample involve administering to the individual an infectious dose of atleast one pathogen that is pathogenic in the specific organ or tissue,such as an attenuated pathogen; and administering to the individual ananti-microbial composition comprising antigenic determinants, theantigenic determinants selected or formulated so that together they arespecific for the at least one pathogen, such as a composition comprisingkilled whole pathogens, so as to treat or prevent an infection by aheterologous micro-organism. The method may involve these twoadministration steps occurring simultaneously. The method may involvethe second step occurring between 1 hour and 30 days after the firststep.

Compositions of the invention may for example be formulated or used foradministration at a site that is distinct from the specific organ ortissue that is targeted for treatment, for example by subcutaneousinjection or intradermal injection. Compositions may for example beformulated for repeated administration, for example by subcutaneous orintradermal injection. In selected embodiments, compositions of theinvention may be formulated or used so as to produce a localized immuneresponse at a site of administration, for example at a site of injectionin the skin.

In selected embodiments, pathogens may be selected for use in methodsand compositions of the invention on the basis that the pathogen isendogenous to the specific organ or tissue that is targeted fortreatment. Alternatively, the pathogen may be exogenous to the specificorgan or tissue. The pathogen may be formulated as an attenuated or akilled pathogen, for example to provide an antigenic composition ofwhole attenuated or killed pathogens. For example, the pathogen may be abacterium, a virus, a protozoa, a fungus, or a helminth.

In a further aspect, a method of formulating an anti-microbialcomposition for treating a condition characterized by an infection in aspecific organ or tissue is provided. The method involves selecting atleast one pathogen that is pathogenic in the specific organ or tissue;producing an antigenic composition comprising antigenic determinantsthat together are specific for the pathogen; and formulating theantigenic composition for administration as an anti-microbialcomposition capable of eliciting an anti-microbial response in thespecific organ or tissue to a heterologous micro-organism.

The method may further involve a diagnostic step of identifying thespecific organ or tissue within which the infection is symptomatic priorto producing the antigenic composition.

Optionally, the antigenic composition may be formulated for subcutaneousinjection or intradermal injection. Optionally, the antigeniccomposition may be formulated for injection to produce a localized skinimmune response at a site of administration. Optionally, the methoddetailed herein is provided such that when a specific tissue or organ isdetermined, the pathogen is selected from a particular group ofpathogens as described herein. In one aspect the pathogen is one that isan endogenous organism, which is a natural cause of infection in thetissue or organ in question. Optionally, the pathogen is an exogenousorganism that is a natural cause of infection in the tissue or organ isquestion, and may include, for example, a bacteria, virus, helminth, orfungus.

Optionally, the antigenic composition may be formulated for repeatedsubcutaneous or intradermal administration. Optionally, the antigeniccomposition may be formulated for administration by a route that is notenteric. Optionally, the pathogen detailed herein is a bacteria, avirus, a protozoa, a fungus or a helminth. Further, the method mayinvolve killing or attenuating the pathogen to formulate the antigeniccomposition as a whole killed or attenuated pathogen composition. Thepathogen may be a member of a species of the endogenous flora that is anatural cause of infection in the specific organ or tissue. The pathogenmay be an exogenous species that is a natural cause of infection in thespecific organ or tissue.

In another aspect, a method of treating an individual for a conditioncharacterized by infection, or a pathology associated with a microbialinfection, in a specific organ or tissue is provided. The methodinvolves administering to the individual an anti-microbial compositioncomprising antigenic determinants. The antigenic determinants areselected or formulated so that together they are specific for at leastone pathogen that is pathogenic in the specific organ or tissue.Optionally, the anti-microbial composition may be administered at anadministration site in successive doses given at a dosage interval ofbetween one hour and one month, over a dosage duration of at least twoweeks. Further, and without limitation, the dosing may comprise two ormore doses (or 10 or more, or 100 or more) over a period from, forexample, 1, 2, 3, 4, 5 or 6 days to 1, 2, 3, 4, 5, or 6 weeks.

In another aspect, use of an anti-microbial composition for treating anindividual for a condition characterized by inflammation in a specificorgan or tissue is disclosed. The anti-microbial composition may forexample contain antigenic determinants selected or formulated so thattogether they are specific for at least one microbial pathogen that ispathogenic in the specific organ or tissue.

In another aspect, use of an anti-microbial composition to formulate amedicament for treating an individual for a condition characterized bypathologies associated with an infection in a specific organ or tissueis disclosed. The anti-microbial composition may for example containantigenic determinants selected or formulated so that together they arespecific for at least one microbial pathogen that is pathogenic in thespecific organ or tissue.

In one aspect, a method of comparing immune responses is provided. Themethod involves administering to an animal having an organ or tissue amedicament having an antigenic composition having antigenic determinantsselected or formulated so that together the antigenic determinants arespecific for at least one microbial pathogen that is pathogenic in theorgan or tissue, extracting a quantifiable immune sample from the organor tissue, measuring a characteristic of the immune response in theorgan or tissue in the quantifiable immune sample following theadministration of the medicament, and, comparing the characteristic ofthe immune response in the quantifiable immune sample to a correspondingcharacteristic of the immune response in a reference immune sampleobtained from a corresponding organ or tissue. Optionally, the referenceimmune sample may be obtained from the corresponding organ or tissue inthe animal prior to the step of administering the medicament.Optionally, the reference immune sample may be obtained from thecorresponding organ or tissue in a second animal. Optionally, the animalmay have an infection situated in the organ or tissue.

Comparing the characteristic of the immune response may involvecomparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells:inflammatory monocytes, macrophages, CD11 b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Further,comparing the characteristic of the immune response may involvecomparing a shift in an activation state of macrophages. Optionally, themacrophages may shift from being M2-like macrophages to being M1-likemacrophages. Further and optionally, the macrophages may shift frombeing M1-like macrophages to being M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cellular markers on any one or more of the following cells: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Themacrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cytokines produced by any one or more of the following cells:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. As detailed herein, the macrophages may include any one or moreof the following: M1-like macrophages or M2-like macrophages.Optionally, the cytokines are produced as a result of a shift in anactivation state of the macrophages. Optionally, the macrophages shiftfrom being M2-like macrophages to being M1-like macrophages. Further andoptionally, the macrophages shift from being M1-like macrophages tobeing M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,differential gene expression produced by any one or more of thefollowing cells: inflammatory monocytes, macrophages, CD11b+ Gr-1+cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ Tcells, or NK cells. The macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Optionally, thedifferential gene expression is produced as a result of a shift in anactivation state of the macrophages. Optionally, macrophages may shiftfrom being M2-like macrophages to being M1-like macrophages. Further andoptionally, the macrophages shift from being M1-like macrophages tobeing M2-like macrophages.

Optionally, the medicament may be administered at an administration sitein successive doses given at a dosage interval of between one hour andone month, over a dosage duration of at least one week. Optionally, themedicament may be administered intradermally or subcutaneously.Optionally, the medicament may be administered in a dose so that eachdose is effective to cause a visible localized inflammatory immuneresponse at the administration site. Optionally, the medicament may beadministered so that visible localized inflammation at theadministration site occurs within 1 to 48 hours. Further and optionally,the animal may be a mammal. Optionally, the animal may be a human or amouse.

In another aspect, a method of selecting a therapeutic preparationsuitable for treating an individual for an infection in a specific organor tissue is provided. The method involves providing an animal having aninfection situated in a specific organ or tissue, providing a testpreparation having one or more antigenic determinants of a microbialpathogen which is pathogenic in the corresponding specific organ ortissue in a healthy individual, measuring a characteristic of the immuneresponse in a reference immune sample obtained from the organ or tissueof the animal, administering the test preparation to the animal,measuring a characteristic of the immune response in a quantifiableimmune sample obtained from a corresponding organ or tissue of theanimal, comparing the characteristic of the immune response in the inthe reference and quantifiable immune samples, and treating an enhancedcharacteristic of the immune response in the quantifiable immune samplecompared to the reference immune sample as an indication of thesuitability of the test preparation as a therapeutic preparation.Optionally, the animal is sacrificed before the quantifiable immunesample has been obtained.

Optionally, comparing the characteristic of the immune response mayinvolve comparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Optionally,comparing the characteristic of the immune response may involvecomparing a shift in an activation state of macrophages. Optionally, themacrophages may shift from being M2-like macrophages to being M1-likemacrophages. Further and optionally, the macrophages may shift frombeing M1-like macrophages to being M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cellular markers on any one or more of the following cells: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Optionally,the macrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cytokines produced by any one or more of the following cells:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages. Optionally, the cytokinesare produced as a result of a shift in an activate state of themacrophages. Optionally, the macrophages may shift from being M2-likemacrophages to being M1-like macrophages. Further, the macrophages mayshift from being M1-like macrophages to being M2-like macrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, differential gene expression produced by any one or moreof the following cells: inflammatory monocytes, macrophages, CD11b+Gr-1+ cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells,CD8+ T cells, or NK cells. Optionally, the macrophages may include anyone or more of the following: M1-like macrophages or M2-likemacrophages. Optionally, the differential gene expression may beproduced as a result of a shift in an activation state of themacrophages. Optionally, the macrophages may shift from being M2-likemacrophages to being M1-like macrophages. Further and optionally, themacrophages may shift from being M1-like macrophages to being M2-likemacrophages.

In another aspect, a method of selectively targeting an immune responseto an infected tissue or an organ in a human subject is provided. Themethod involves administering to the subject a medicament having aneffective amount of a microbial pathogen antigenic composition, whereinthe microbial pathogen may be pathogenic in the specific organ or tissueof the subject in which there is an infection caused by a heterologousmicro-organism, and the antigenic composition comprises antigenicdeterminants that together are specific for the microbial pathogen.Optionally, the antigenic composition may include a whole killedbacterial cell composition. Optionally, the medicament may beadministered to the subject in an amount and for a time that iseffective to up-regulate an anti-microbial immune response in the organor tissue of the subject in which there is an infection caused by aheterologous micro-organism. Optionally, the method may further involvemeasuring a characteristic of the immune response. The method alsoincludes prophylactic treatment of infections, by immune-protectivevaccination.

In another aspect, a method for treating a human subject for aninfection, or a pathology associated with a microbial infection,situated in a tissue or an organ is provided. The method involvesadministering to the subject a medicament having an effective amount ofan antigenic composition comprising a microbial pathogen, such as wholekilled bacterial cell or viral compositions, wherein the microbialpathogen is pathogenic in the specific organ or tissue of the subjectwithin which the heterologous microbial infection is situated or withinwhich the future infection is to be prevented. The medicament may beadministered to the subject in an amount and for a time that iseffective to modulate an immune response in the target organ or tissue.In this way, the invention provides site specific immunomodulators(SSTs), which elicit an immunological response in a target organ ortissue. In select embodiments, the target organ or tissue may bedistinct or distant from the site of administration. Optionally, themodulation of the immune response may involve a shift in the activationstate of macrophages. Optionally, the modulation of the immune responsemay involve shifting from a M2-like macrophage response to a M1-likemacrophage response. The modulation of the immune response may involve ashift from M1-like macrophages to M2-like macrophages, as those termsare defined herein. Optionally and without limitation, the method mayfurther involve measuring a characteristic of the immune response.

Optionally, comparing the characteristic of the immune response mayinvolve comparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Optionally,comparing the characteristic of the immune response may involvecomparing a shift in an activation state of macrophages. Further andoptionally, the macrophages may shift from being M2-like macrophages tobeing M1-like macrophages. Optionally, the macrophages may shift frombeing M1-like macrophages to being M2-like macrophages.

Further and without limitation, comparing the characteristic of theimmune response may involve identifying, in the quantifiable andreference immune samples, cellular markers on any one or more of thefollowing cells: inflammatory monocytes, macrophages, CD11b+ Gr-1+cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ Tcells, or NK cells. The macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Optionally,comparing the characteristic of the immune response may involveidentifying, in the quantifiable and reference immune samples, cytokinesproduced by any one or more of the following cells: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Optionally,the macrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages. Further, cytokines may be producedas a result of a shift in an activation state of the macrophages. Themacrophages may shift from being M2-like macrophages to being M1-likemacrophages. Optionally, the macrophages may shift from being M1-likemacrophages to being M2-like macrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, differential gene expression produced by any one or moreof the following cells: inflammatory monocytes, macrophages, CD11b+Gr-1+ cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells,CD8+ T cells, or NK cells. The macrophages may include any one or moreof the following: M1-like macrophages or M2-like macrophages.Optionally, the differential gene expression may be produced as a resultof a shift in an activation state of the macrophages. Further andoptionally, the macrophages may shift from being M2-like macrophages tobeing M1-like macrophages. The macrophages may shift from being M1-likemacrophages to being M2-like macrophages.

In another aspect, a method of monitoring efficacy of a treatment regimein an individual being treated for an infection in a specific organ ortissue is provided. The method involves measuring a characteristic of animmune response in a post-treatment immune sample obtained from thespecific organ or tissue after the individual has been subject to thetreatment regime for a period of time, wherein the presence of acharacteristic of the immune response which is greater in magnitude thanwould be expected had the individual not been subject to the treatmentregime, is indicative of the efficacy of the treatment regime; and thetreatment regime involves administering a preparation comprising one ormore antigenic determinants of a microbial pathogen which is pathogenicin the corresponding specific organ or tissue in a healthy subject.

The method detailed herein may further involve measuring thecharacteristic of the immune response in a pre-treatment referencesample, wherein the pre-treatment reference sample was obtained from thespecific organ or tissue before, at the same time as or aftercommencement of the treatment regime, but prior to obtaining thepost-treatment immune sample, and comparing the characteristic of theimmune response in the pre-treatment and post-treatment samples, whereinan increase in the magnitude of the immune response in thepost-treatment immune sample compared to the pre-treatment referencesample is indicative of the efficacy of the treatment regime.Optionally, measuring the characteristic of the immune response mayinvolve determining an indication of the number of inflammatorymonocytes in a sample of the organ or tissue. Optionally, measuring thecharacteristic of the immune response may involve determining anindication of the number of macrophages in a sample of the organ ortissue. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages.

As detailed herein in another aspect, the invention also providesmethods for formulating an immunogenic composition for treating aninfection situated in a specific organ or tissue in a mammal, such ashuman patient. The method may include selecting at least one microbialpathogen that is naturally pathogenic in the organ or tissue of themammal within which the heterologous microbial infection is situated. Anantigenic composition may be produced that includes antigenicdeterminants that together are specific for or characteristic of themicrobial pathogen.

A diagnostic step may be used to identify the specific organ or tissuewithin which the infection is situated, prior to producing the antigeniccomposition targeted to the site of the infection. The site of theinfection may be a primary site, or a secondary site of metastasis. Theantigenic composition may be sufficiently specific that it would becapable of eliciting an immune response in the mammal specific to themicrobial pathogen. The antigenic composition may be a bacterialcomposition, for example derived from a bacterial species or speciesthat are endogenous to the flora of the patient or from an exogenousspecies or species. In alternative embodiments, the antigeniccomposition may be derived from a virus or viruses. Accordingly, themicrobial pathogen from which the antigenic composition is derived maybe a virus. The microbial pathogen may be killed. In alternativeembodiments, the microbial pathogen may be live or attenuated.Immunogenic compositions of the invention may also be formulated oradministered with anti-microbial modalities, such as an NSAID. The siteof administration may be at a site distant from the site of theinfection, for example in an organ or tissue that is not the organ ortissue within which the heterologous infection is situated, for examplethe skin or subcutaneous tissue.

The antigenic composition may for example be formulated for subcutaneousinjection, intradermal injection or oral administration. In embodimentsfor subcutaneous or intradermal injection, the dosing or formulation ofthe antigenic composition may be adjusted in order to produce alocalized immune reaction visible in the skin at the site ofadministration, for example an area of inflammation from 2 mm to 100 mmin diameter appearing, for example, 2-48 hours after administration andlasting, for example, 2-72 hours or longer. The antigenic compositionmay be formulated for repeated subcutaneous or intradermaladministration, for example at alternating successive sites.

In some embodiments, the invention involves methods of treating a mammalfor an infection situated in a tissue or an organ. In alternativeembodiments, the treatment may anticipate the development of theheterologous infection in the tissue, for example if the site of aprimary infection suggests the likelihood of the spread of the infectionto a particular tissue or organ, then the patient may beprophylactically treated to prevent or ameliorate metastasis to thattissue or organ. The method may include administering to the subject aneffective amount of an antigenic composition comprising antigenicdeterminants that together are specific for at least one microbialpathogen. An aspect of the invention involves the use of a microbialpathogen that is pathogenic in the specific organ or tissue of themammal within which the heterologous infection is situated. Theantigenic composition may be administered, for example by subcutaneousor intradermal injection at an administration site, in successive dosesgiven at a dosage interval, for example of between one hour and onemonth, over a dosage duration, for example of at least 1 week, 2 weeks,2 months, 6 months, 1, 2, 3, 4, or 5 years or longer. Each injectiondose may for example be metered so that it is effective to cause visiblelocalized inflammation at the administration site, appearing, forexample, 1-48 hours after injection.

In another aspect, methods are provided for treating heterologousinfections of a specific organ or tissue in a subject by administeringone or more antigens of one or more microbial pathogens, such asbacterial, viral or fungal species that are pathogenic in the specificorgan or tissue.

In alternative embodiments, the pathogenic microbial species may becapable of causing infection naturally, (i.e., without humanintervention) in the specific organ or tissue in a healthy subject, ormay have caused an infection in the specific organ or tissue in ahealthy subject. In alternative embodiments, the antigen may beadministered by administering preparations derived from whole cells of amicrobial species. In alternative embodiments, the method may, forexample, include administering at least two or more microbial species,or administering at least three or more microbial species, and themicrobes may be bacteria or viruses. In alternative embodiments, themethod may further include administering a supplement or an adjuvant. Anaspect of the invention involves administering antigenic compositions soas to elicit an immune response in said subject.

In alternative embodiments, the microbial pathogen in the antigeniccomposition may be killed, and thus rendered non-infectious. In someembodiments, the antigenic composition is administered at a site distantfrom the heterologous infection site, and in selected embodiments ofthis kind, methods of the invention may be carried out so that they donot produce infection at the heterologous infection site.

As detailed herein, various aspects of the invention involve treatingheterologous infections. In this context, treatment may be carried outso as to provide a variety of outcomes. For example, treatment may:provoke an immune reaction that is effective to inhibit or amelioratethe growth or proliferation of an infection; inhibit the growth orproliferation of heterologous micro-organisms; cause remission of aninfection; improve quality of life; reduce the risk of recurrence of aninfection; inhibit spread of an infection; or, improve patient survivalrates in a patient population. In this context, extending the lifeexpectancy of a patient, or patient population, means to increase thenumber of patients who survive for a given period of time following aparticular diagnosis. In some embodiments, treatment may be of patientswho have not responded to other treatments, such as patients for whom achemotherapy or surgery has not been an effective treatment. Treatmentin alternative embodiments may for example be before or after onset ofheterologous infection. For example prophylactic treatment may beundertaken, for example of patients diagnosed as being at risk of aparticular heterologous infection.

Bacteria and Bacterial Colonizations and Infections

Most animals are colonized to some degree by other organisms, such asbacteria, which generally exist in symbiotic or commensal relationshipswith the host animal. Thus, many species of normally harmless bacteriaare found in healthy animals, and are usually localized to the surfaceof specific organs and tissues. Often, these bacteria aid in the normalfunctioning of the body. For example, in humans, symbiotic Escherichiacoli bacteria may be found in the intestine, where they promote immunityand reduce the risk of infection with more virulent pathogens.

Bacteria that are generally harmless, such as Escherichia coli, cancause infection in healthy subjects, with results ranging from mild tosevere infection to death. Whether or not a bacterium is pathogenic(i.e., causes infection) depends to some extent on factors such as theroute of entry and access to specific host cells, tissues, or organs;the intrinsic virulence of the bacterium; the amount of the bacteriapresent at the site of potential infection; or the health of the hostanimal. Thus, bacteria that are normally harmless can become pathogenicgiven favorable conditions for infection, and even the most virulentbacterium requires specific circumstances to cause infection.Accordingly, microbial species that are members of the normal flora canbe pathogens when they move beyond their normal ecological role in theendogenous flora. For example, endogenous species can cause infectionoutside of their ecological niche in regions of anatomical proximity,for example by contiguous spread. When this occurs, these normallyharmless endogenous bacteria are considered pathogenic.

Specific bacterial species and viruses are known to cause infections inspecific cells, tissues, or organs in otherwise healthy subjects.Examples of bacteria and viruses that commonly cause infections inspecific organs and tissues of the body are listed below; it will beunderstood that these examples are not intended to be limiting and thata skilled person would be able to readily recognize and identifyinfectious or pathogenic bacteria that cause infections, or commonlycause infections, in various organs and tissues in healthy adults (andrecognize the relative frequency of infection with each bacterialspecies) based on the knowledge in the field as represented, forexample, by the following publications: Manual of Clinical Microbiology8th Edition, Patrick Murray, Ed., 2003, ASM Press American Society forMicrobiology, Washington D.C., USA; Mandell, Douglas, and Bennett'sPrinciples and Practice of Infectious Diseases 5th Edition, G. L.Mandell, J. E. Bennett, R. Dolin, Eds., 2000, Churchill Livingstone,Philadelphia, Pa., USA, all of which are incorporated by referenceherein.

Infections of the skin are commonly caused by the following bacterialspecies: Staphylococcus aureus, Beta hemolytic streptococci group A, B,C or G, Corynebacterium diptheriae, Corynebacterium ulcerans, orPseudomonas aeruginosa; or viral pathogens: rubeola, rubella,varicella-zoster, echoviruses, coxsackieviruses, adenovirus, vaccinia,herpes simplex, or parvo B19.

Infections of the soft tissue (e.g., fat and muscle) are commonly causedby the following bacterial species: Streptococcus pyogenes,Staphylococcus aureus, Clostridium perfringens, or other Clostridiumspp.; or viral pathogens: influenza, or coxsackieviruses.

Infections of the breast are commonly caused by the following bacterialspecies: Staphylococcus aureus, or Streptococcus pyogenes.

Infections of the lymph nodes of the head and neck are commonly causedby the following bacterial species: Staphylococcus aureus, orStreptococcus pyogenes; or viral pathogens: Epstein-Barr,cytomegalovirus, adenovirus, measles, rubella, herpes simplex,coxsackieviruses, or varicella-zoster.

Infections of the lymph nodes of the arm/axillae are commonly caused bythe following bacterial species: Staphylococcus aureus, or Streptococcuspyogenes; or viral pathogens: measles, rubella, Epstein-Barr,cytomegalovirus, adenovirus, or varicella-zoster.

Infections of the lymph nodes of the mediastinum are commonly caused bythe following bacterial species: viridans streptococci, Peptococcusspp., Peptostreptococcus spp., Bacteroides spp., Fusobacterium spp., orMycobacterium tuberculosis; or viral pathogens: measles, rubella,Epstein-Barr, cytomegalovirus, varicella-zoster, or adenovirus.

Infections of the pulmonary hilar lymph nodes are commonly caused by thefollowing bacterial species: Streptococcus pneumoniae, Moraxellacatarrhalis, Mycoplasma pneumoniae, Klebsiella pneumoniae, Haemophilusinfluenza, Chlamydophila pneumoniae, Bordetella pertussis orMycobacterium tuberculosis; or viral pathogens: influenza, adenovirus,rhinovirus, coronavirus, parainfluenza, respiratory syncytial virus,human metapneumovirus, or coxsackievirus.

Infections of the intra-abdominal lymph nodes are commonly caused by thefollowing bacterial species: Yersinia enterocolitica, Yersiniapseudotuberculosis, Salmonella spp., Streptococcus pyogenes, Escherichiacoli, Staphylococcus aureus, or Mycobacterium tuberculosis; or viralpathogens: measles, rubella, Epstein-Barr, cytomegalovirus,varicella-zoster, adenovirus, influenza, or coxsackieviruses.

Infections of the lymph nodes of the leg/inguinal region are commonlycaused by the following bacterial species: Staphylococcus aureus, orStreptococcus pyogenes; or viral pathogens: measles, rubella,Epstein-Barr, cytomegalovirus, or herpes simplex.

Infections of the blood (i.e., septicemia) are commonly caused by thefollowing bacterial species: Staphylococcus aureus, Streptococcuspyogenes, coagulase-negative staphylococci, Enterococcus spp.,Escherichia coli, Klebsiella spp., Enterobacter spp., Proteus spp.,Pseudomonas aeruginosa, Bacteroides fragilis, Streptococcus pneumoniae,or group B streptococci; or viral pathogens: rubeola, rubella,varicella-zoster, echoviruses, coxsackieviruses, adenovirus,Epstein-Barr, herpes simplex, or cytomegalovirus.

Infections of the bone are commonly caused by the following bacterialspecies: Staphylococcus aureus, coagulase-negative staphylococci,Streptococcus pyogenes, Streptococcus pneumoniae, Streptococcusagalactiae, other streptococci spp., Escherichia coli, Pseudomonas spp.,Enterobacter spp., Proteus spp., or Serratia spp.; or viral pathogens:parvovirus B19, rubella, or hepatitis B.

Infections of the joint are commonly caused by the following bacterialspecies: Staphylococcus aureus, coagulase-negative staphylococci,Streptococcus pyogenes, Streptococcus pneumoniae, Streptococcusagalactiae, other streptococci spp., Escherichia coli, Pseudomonas spp.,Enterobacter spp., Proteus spp., Serratia spp., Neisseria gonorrhea,salmonella species, Mycobacterim tuberculosis, Hemophilus influenza; orviral pathogens: parvovirus B19, rubella, hepatitis B; or fungalpathogen: Scedosporium prolificans

Infections of the meninges are commonly caused by the followingbacterial species: Haemophilus influenzae, Neisseria meningitidis,Streptococcus pneumoniae, Streptococcus agalactiae, or Listeriamonocytogenes; or viral pathogens: echoviruses, coxsackieviruses, otherenteroviruses, or mumps.

Infections of the brain are commonly caused by the following bacterialspecies: Streptococcus spp. (including S. anginosus, S. constellatus, S.intermedius), Staphylococcus aureus, Bacteroides spp., Prevotella spp.,Proteus spp., Escherichia coli, Klebsiella spp., Pseudomonas spp.,Enterobacter spp., or Borrelia burgdorferi; or viral pathogens:coxsackieviruses, echoviruses, poliovirus, other enteroviruses, mumps,herpes simplex, varicella-zoster, flaviviruses, or bunyaviruses.

Infections of the spinal cord are commonly caused by the followingbacterial species: Haemophilus influenzae, Neisseria meningitidis,Streptococcus pneumoniae, Streptococcus agalactiae, Listeriamonocytogenes, or Borrelia burgdorferi; or viral pathogens:coxsackieviruses, echoviruses, poliovirus, other enteroviruses, mumps,herpes simplex, varicella-zoster, flaviviruses, or bunyaviruses.

Infections of the eye/orbit are commonly caused by the followingbacterial species: Staphylococcus aureus, Streptococcus pyogenes,Streptococcus pneumoniae, Streptococcus milleri, Escherichia coli,Bacillus cereus, Chlamydia trachomatis, Haemophilus influenza,Pseudomonas spp., Klebsiella spp., or Treponema pallidum; or viralpathogens: adenoviruses, herpes simplex, varicella-zoster, orcytomegalovirus.

Infections of the salivary glands are commonly caused by the followingbacterial species: Staphylococcus aureus, viridans streptococci (e.g.,Streptococcus salivarius, Streptococcus sanguis, Streptococcus mutans),Peptostreptococcus spp., or Bacteroides spp., or other oral anaerobes;or viral pathogens: mumps, influenza, enteroviruses, or rabies.

Infections of the mouth are commonly caused by the following bacterialspecies: Prevotella melaninogenicus, anaerobic streptococci, viridansstreptococci, Actinomyces spp., Peptostreptococcus spp., or Bacteroidesspp., or other oral anaerobes; or viral pathogens: herpes simplex,coxsackieviruses, or Epstein-Barr.

Infections of the tonsils are commonly caused by the following bacterialspecies: Streptococcus pyogenes, or Group C or G B-hemolyticstreptococci; or viral pathogens: rhinoviruses, influenza, coronavirus,adenovirus, parainfluenza, respiratory syncytial virus, or herpessimplex.

Infections of the sinuses are commonly caused by the following bacterialspecies: Streptococcus pneumoniae, Haemophilus influenza, Moraxellacatarrhalis, a-streptococci, anaerobic bacteria (e.g., Prevotella spp.),or Staphylococcus aureus; or viral pathogens: rhinoviruses, influenza,adenovirus, or parainfluenza.

Infections of the nasopharynx are commonly caused by the followingbacterial species: Streptococcus pyogenes, or Group C or G B-hemolyticstreptococci; or viral pathogens: rhinoviruses, influenza, coronavirus,adenovirus, parainfluenza, respiratory syncytial virus, or herpessimplex.

Infections of the thyroid are commonly caused by the following bacterialspecies: Staphylococcus aureus, Streptococcus pyogenes, or Streptococcuspneumoniae; or viral pathogens: mumps, or influenza.

Infections of the larynx are commonly caused by the following bacterialspecies: Mycoplasma pneumoniae, Chlamydophila pneumoniae, orStreptococcus pyogenes; or viral pathogens: rhinovirus, influenza,parainfluenza, adenovirus, corona virus, or human metapneumovirus.

Infections of the trachea are commonly caused by the following bacterialspecies: Mycoplasma pneumoniae; or viral pathogens: parainfluenza,influenza, respiratory syncytial virus, or adenovirus.

Infections of the bronchi are commonly caused by the following bacterialspecies: Mycoplasma pneumoniae, Chlamydophila pneumoniae, Bordetellapertussis, Streptococcus pneumoniae, or Haemophilus influenzae; or viralpathogens: influenza, adenovirus, rhinovirus, coronavirus,parainfluenza, respiratory syncytial virus, human metapneumovirus, orcoxsackievirus.

Infections of the lung are commonly caused by the following bacterialspecies: Streptococcus pneumoniae, Moraxella catarrhalis, Mycoplasmapneumoniae, Klebsiella pneumoniae, or Haemophilus influenza; or viralpathogens: influenza, adenovirus, respiratory syncytial virus, orparainfluenza.

Infections of the pleura are commonly caused by the following bacterialspecies: Staphylococcus aureus, Streptococcus pyogenes, Streptococcuspneumoniae, Haemophilus influenzae, Bacteroides fragilis, Prevotellaspp., Fusobacterium nucleatum, peptostreptococcus spp., or Mycobacteriumtuberculosis; or viral pathogens: influenza, adenovirus, respiratorysyncytial virus, or parainfluenza.

Infections of the mediastinum are commonly caused by the followingbacterial species: viridans streptococci, Peptococcus spp.,Peptostreptococcus spp., Bacteroides spp., Fusobacterium spp., orMycobacterium tuberculosis; or viral pathogens: measles, rubella,Epstein-Barr, or cytomegalovirus.

Infections of the heart are commonly caused by the following bacterialspecies: Streptococcus spp. (including S. mitior, S. bovis, S. sanguis,S. mutans, S. anginosus), Enterococcus spp., Staphylococcus spp.,Corynebacterium diptheriae, Clostridium perfringens, Neisseriameningitidis, or Salmonella spp.; or viral pathogens: enteroviruses,coxsackieviruses, echoviruses, poliovirus, adenovirus, mumps, rubeola,or influenza.

Infections of the esophagus are commonly caused by the followingbacterial species: Actinomyces spp., Mycobacterium avium, Mycobacteriumtuberculosis, or Streptococcus spp.; or viral pathogens:cytomegalovirus, herpes simplex, or varicella-zoster.

Infections of the stomach are commonly caused by the following bacterialspecies: Streptococcus pyogenes or Helicobacter pylori; or viralpathogens: cytomegalovirus, herpes simplex, Epstein-Barr, rotaviruses,noroviruses, or adenoviruses.

Infections of the small bowel are commonly caused by the followingbacterial species: Escherichia coli, Clostridium difficile, Bacteroidesfragilis, Bacteroides vulgatus, Bacteroides thetaiotaomicron,Clostridium perfringens, Salmonella enteriditis, Yersiniaenterocolitica, or Shigella flexneri; or viral pathogens: adenoviruses,astroviruses, caliciviruses, noroviruses, rotaviruses, orcytomegalovirus.

Infections of the colon/rectum are commonly caused by the followingbacterial species: Escherichia coli, Clostridium difficile, Bacteroidesfragilis, Bacteroides vulgatus, Bacteroides thetaiotaomicron,Clostridium perfringens, Salmonella enteriditis, Yersiniaenterocolitica, or Shigella flexneri; or viral pathogens: adenoviruses,astroviruses, caliciviruses, noroviruses, rotaviruses, orcytomegalovirus.

Infections of the anus are commonly caused by the following bacterialspecies: Streptococcus pyogenes, Bacteroides spp., Fusobacterium spp.,anaerobic streptococci, Clostridium spp., Escherichia coli, Enterobacterspp., Pseudomonas aeruginosa, or Treponema pallidum; or viral pathogens:herpes simplex.

Infections of the perineum are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella spp., Enterococcus spp.,Bacteroides spp., Fusobacterium spp., Clostridium spp., Pseudomonasaeruginosa, anaerobic streptococci, Clostridium spp., or Enterobacterspp.; or viral pathogens: herpes simplex.

Infections of the liver are commonly caused by the following bacterialspecies: Escherichia coli, Klebsiella spp., Streptococcus (anginosusgroup), Enterococcus, spp. other viridans streptococci, or Bacteroidesspp.; or viral pathogens: hepatitis A, Epstein-Barr, herpes simplex,mumps, rubella, rubeola, varicella-zoster, coxsackieviruses, oradenovirus.

Infections of the gallbladder are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella spp., Enterobacter spp.,enterococci, Bacteroides spp., Fusobacterium spp., Clostridium spp.,Salmonella enteriditis, Yersinia enterocolitica, or Shigella flexneri.

Infections of the biliary tract are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella spp., Enterobacter spp.,enterococci, Bacteroides spp., Fusobacterium spp., Clostridium spp.,Salmonella enteriditis, Yersinia enterocolitica, or Shigella flexneri;or viral pathogens: hepatitis A, Epstein-Barr, herpes simplex, mumps,rubella, rubeola, varicella-zoster, cocsackieviruses, or adenovirus.

Infections of the pancreas are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella spp., Enterococcus spp.,Pseudomonas spp., Staphylococcal spp., Mycoplasma spp., Salmonellatyphi, Leptospirosis spp., or Legionella spp.; or viral pathogens:mumps, coxsackievirus, hepatitis B, cytomegalovirus, herpes simplex 2,or varicella-zoster.

Infections of the spleen are commonly caused by the following bacterialspecies: Streptococcus spp., Staphylococcus spp., Salmonella spp.,Pseudomonas spp., Escherichia coli, or Enterococcus spp.; or viralpathogens: Epstein-Barr, cytomegalovirus, adenovirus, measles, rubella,coxsackieviruses, or varicella-zoster.

Infections of the adrenal gland are commonly caused by the followingbacterial species: Streptococcus spp., Staphylococcus spp., Salmonellaspp., Pseudomonas spp., Escherichia coli, or Enterococcus spp.; or viralpathogens: varicella-zoster.

Infections of the kidney are commonly caused by the following bacterialspecies: Escherichia coli, Proteus mirabilis, Proteus vulgatus,Providentia spp., Morganella spp., Enterococcus faecalis, or Pseudomonasaeruginosa; or viral pathogens: BK virus, or mumps.

Infections of the ureter are commonly caused by the following bacterialspecies: Escherichia coli, Proteus mirabilis, Proteus vulgatus,Providentia spp., Morganella spp., or Enterococcus spp.

Infections of the bladder are commonly caused by the following bacterialspecies: Escherichia coli, Proteus mirabilis, Proteus vulgatus,Providentia spp., Morganella spp., Enterococcus faecalis, orCorynebacterium jekeum; or viral pathogens: adenovirus, orcytomegalovirus.

Infections of the peritoneum are commonly caused by the followingbacterial species: Staphylococcus aureus, Streptococcus pyogenes,Streptococcus pneumonia, Escherichia coli, Klebsiella spp., Proteusspp., enterococci, Bacteroides fragilis, Prevotella melaninogenica,Peptococcus spp., Peptostreptococcus spp., Fusobacterium spp., orClostridium spp.

Infections of the retroperitoneal area are commonly caused by thefollowing bacterial species: Escherichia coli, or Staphylococcus aureus.

Infections of the prostate are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella spp., Enterobacter spp.,Proteus mirabilis, enterococci spp., Pseudomonas spp., Corynebacteriumspp., or Neisseria gonorrhoeae; or viral pathogens: herpes simplex.

Infections of the testicle are commonly caused by the followingbacterial species: Escherichia coli, Klebsiella pneumoniae, Pseudomonasaeruginosa, Staphylococcus spp., Streptococcus spp., or Salmonellaenteriditis; or viral pathogens: mumps, coxsackievirus, or lymphocyticchoriomeningitis virus.

Infections of the penis are commonly caused by the following bacterialspecies: Staphylococcus aureus, Streptococcus pyogenes, Neisseriagonorrhoeae, or Treponema pallidum; or viral pathogens: herpes simplex.

Infections of the ovary/adnexae are commonly caused by the followingbacterial species: Neisseria gonorrhoeae, Chlamydia trachomatis,Gardenerella vaginalis, Prevotella spp., Bacteroides spp., Peptococcusspp. Streptococcus spp., or Escherichia coli.

Infections of the uterus are commonly caused by the following bacterialspecies: Neisseria gonorrhoeae, Chlamydia trachomatis, Gardenerellavaginalis, Prevotella spp., Bacteroides spp., Peptococcus spp.,Streptococcus spp., or Escherichia coli.

Infections of the cervix are commonly caused by the following bacterialspecies: Neisseria gonorrhoeae, Chlamydia trachomatis, or Treponemapallidum; or viral pathogens: herpes simplex.

Infections of the vagina are commonly caused by the following bacterialspecies: Gardenerella vaginalis, Prevotella spp., Bacteroides spp.,peptococci spp., Escherichia coli, Neisseria gonorrhoeae, ChlamydiaTrachomatis, or Treponema pallidum; or viral pathogens: herpes simplex.

Infections of the vulva are commonly caused by the following bacterialspecies: Staphylococcus aureus, Streptococcus pyogenes, or Treponemapallidum; or viral pathogens: herpes simplex.

Bacterial Strains/Viral Subtypes

It will be understood by a skilled person in the art that bacterialspecies are classified operationally as collections of similar strains(which generally refers to groups of presumed common ancestry withidentifiable physiological but usually not morphological distinctions,and which may be identified using serological techniques againstbacterial surface antigens). Thus, each bacterial species (e.g.,Streptococcus pneumoniae) has numerous strains (or serotypes), which maydiffer in their ability to cause infection or differ in their ability tocause infection in a particular organ/site. For example, although thereare at least 90 serotypes of Streptococcus pneumoniae, serotypes 1, 3,4, 7, 8, and 12 are most frequently responsible for pneumococcal diseasein humans.

As a second example, certain strains of Escherichia coli, referred to asextraintestinal pathogenic E. coli (ExPEC), are more likely to causeurinary tract infection or other extraintestinal infections such asneonatal meningitis, whereas other strains, including enterotoxigenic E.coli (ETEC), enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli(EHEC), Shiga toxin-producing E. coli (STEC), enteroaggregative E. coli(EAEC), enteroinvasive E. coli (EIEC) and diffuse adhering E. coli(DAEC) are more likely to cause gastrointestinal infection/diarrhea.Even among the sub-category of ExPEC strains, specific virulence factors(e.g., production of type-1 fimbriae) enable certain strains to be morecapable of causing infection of the bladder, while other virulencefactors (e.g., production of P fimbriae) enable other strains to be morecapable of causing infection in the kidneys. In accordance with thepresent invention, an ExPEC strain(s) that is more likely to causeinfection in the bladder may be chosen for a formulation to targetbladder infection, whereas an ExPEC strain(s) that is more likely tocause infection in the kidney may be chosen for a formulation to targetkidney infection. Likewise, one or more of an ETEC, EPEC, EHEC, STEC,EAEC, EIEC or DAEC strains of E. coli (i.e., strains that cause coloninfection), may be chosen for a formulation to treat colon infections.

Similarly, there may be numerous subtypes of specific viruses. Forexample, there are three types of influenza viruses, influenza A,influenza B and influenza C, which differ in epidemiology, host rangeand clinical characteristics. For example, influenza A is more likely tobe associated with viral lung infection, whereas influenza B is morelikely to be associated with myositis (i.e., muscle infection).Furthermore, each of these three types of influenza virus have numeroussubtypes, which also may differ in epidemiology, host range and clinicalcharacteristics. In accordance with the present invention, one maychoose an influenza A subtype most commonly associated with lunginfection to target heterologous lung infections, whereas one may choosean influenza B strain most commonly associated with myositis to treatinfections of the muscle/soft tissues.

It is understood that a clinical microbiologist skilled in the art wouldtherefore be able to select, based on the present disclosure and thebody of art relating to bacterial strains for each species of bacteria(and viral subtypes for each type of virus), the strains of a particularbacterial species (or subtype of a particular virus) to target aspecific organ or tissue. In this way, the invention provides sitespecific immunomodulators (SSIs), in the sense that the formulations andtreatments of the invention elicit an immunological response in a targetorgan or tissue, and that target may be distinct or distant from thesite of administration.

Microbial Compositions, Dosages, and Administration

The compositions of the invention include antigens of pathogenicmicrobial (bacterial or viral) species that are pathogenic in a specifictissue or organ. The compositions may include whole bacterial species,or may include extracts or preparations of the pathogenic bacterialspecies of the invention, such as cell wall or cell membrane extracts,or whole cells, or exotoxins, or whole cells and exotoxins. Thecompositions may also include one or more isolated antigens from one ormore of the pathogenic bacterial species of the invention; in someembodiments, such compositions may be useful in situations where it maybe necessary to precisely administer a specific dose of a particularantigen, or may be useful if administering a whole bacterial species orcomponents thereof (e.g., toxins) may be harmful. Pathogenic bacterialspecies may be available commercially (from, for example, ATCC(Manassas, Va., USA), or may be clinical isolates from subjects having abacterial infection of a tissue or organ (e.g., pneumonia).

The microbial compositions of the invention can be provided alone or incombination with other compounds (for example, nucleic acid molecules,small molecules, peptides, or peptide analogues), in the presence of aliposome, an adjuvant, or any pharmaceutically acceptable carrier, in aform suitable for administration to mammals, for example, humans. Asused herein “pharmaceutically acceptable carrier” or “excipient”includes any and all solvents, dispersion media, coatings, antibacterialand antifungal agents, isotonic and absorption delaying agents, and thelike that are physiologically compatible. The carrier can be suitablefor any appropriate form of administration, including subcutaneous,intradermal, intravenous, parenteral, intraperitoneal, intramuscular,sublingual, inhalational, intratumoral or oral administration.Pharmaceutically acceptable carriers include sterile aqueous solutionsor dispersions and sterile powders for the extemporaneous preparation ofsterile injectable solutions or dispersion. The use of such media andagents for pharmaceutically active substances is well known in the art.Except insofar as any conventional media or agent is incompatible withthe active compound (i.e., the specific bacteria, bacterial antigens, orcompositions thereof of the invention), use thereof in thepharmaceutical compositions of the invention is contemplated.Supplementary active compounds can also be incorporated into thecompositions.

Conventional pharmaceutical practice may be employed to provide suitableformulations or compositions to administer the compounds to subjectssuffering from an infection. Any appropriate route of administration maybe employed, for example, parenteral, intravenous, intradermal,subcutaneous, intramuscular, intracranial, intraorbital, ophthalmic,intraventricular, intracapsular, intraspinal, intrathecal,intracisternal, intraperitoneal, intranasal, inhalational, aerosol,topical, intratumoral, sublingual or oral administration. Therapeuticformulations may be in the form of liquid solutions or suspensions; fororal administration, formulations may be in the form of tablets orcapsules; for intranasal formulations, in the form of powders, nasaldrops, or aerosols; and for sublingual formulations, in the form ofdrops, aerosols or tablets.

Methods well known in the art for making formulations are found in, forexample, “Remington's Pharmaceutical Sciences” (20th edition), ed. A.Gennaro, 2000, Mack Publishing Company, Easton, Pa. Formulations forparenteral administration may, for example, contain excipients, sterilewater, or saline, polyalkylene glycols such as polyethylene glycol, oilsof vegetable origin, or hydrogenated napthalenes. Biocompatible,biodegradable lactide polymer, lactide/glycolide copolymer, orpolyoxyethylene-polyoxypropylene copolymers may be used to control therelease of the compounds. Other potentially useful parenteral deliverysystems for include ethylene-vinyl acetate copolymer particles, osmoticpumps, implantable infusion systems, and liposomes. Formulations forinhalation may contain excipients, for example, lactose, or may beaqueous solutions containing, for example, polyoxyethylene-9-laurylether, glycocholate and deoxycholate, or may be oily solutions foradministration in the form of nasal drops, or as a gel. For therapeuticor prophylactic compositions, the pathogenic bacterial species areadministered to an individual in an amount effective to stop or slowprogression of the infection, or to increase survival of the subject.

An “effective amount” of a pathogenic microbial species or antigenthereof according to the invention includes a therapeutically effectiveamount or a prophylactically effective amount. A “therapeuticallyeffective amount” refers to an amount effective, at dosages and forperiods of time necessary, to achieve the desired therapeutic result,such as reduction or elimination of the heterologous infection,prevention of microbial infection processes, slowing the growth of thetumour, or an increase in survival time beyond that which is expectedusing for example the SEER database. A therapeutically effective amountof a pathogenic microbial (bacterial or viral) species or antigen(s)thereof may vary according to factors such as the disease state, age,sex, and weight of the individual, and the ability of the compound toelicit a desired response in the individual. Dosage regimens may beadjusted to provide the optimum therapeutic response. A therapeuticallyeffective amount may also be one in which any toxic or detrimentaleffects of the pathogenic bacterial species or virus or antigen thereofare outweighed by the therapeutically beneficial effects. A“prophylactically effective amount” refers to an amount effective, atdosages and for periods of time necessary, to achieve the desiredprophylactic result, such as prevention of infection, slowing theprogress of the infection, reduction or elimination of the heterologousmicrobial cells.

For administration by subcutaneous or intradermal injection, anexemplary range for therapeutically or prophylactically effectiveamounts of one or more pathogenic bacterial species may be about 1million to 100,000 million organisms per ml, or may be 100 million to7000 million organisms per ml, or may be 500 million to 6000 millionorganisms per ml, or may be 1000 million to 5000 million organisms perml, or may be 2000 million to 4000 million organisms per ml, or anyinteger within these ranges. The total concentration of bacteria per mlmay range from 1 million to 100,000 million organisms per ml, or may be50 million to 7000 million organisms per ml, or may be 100 million to6000 million organisms per ml, or may be 500 million to 5000 millionorganisms per ml, or may be 1000 million to 4000 million organisms perml, or any integer within these ranges. The range for therapeutically orprophylactically effective amounts of antigens of a pathogenic bacterialspecies may be any integer from 0.1 nM-0.1 M, 0.1 nM-0.05M, 0.05 nM-15μM or 0.01 nM-10 μM.

It is to be noted that dosage concentrations and ranges may vary withthe severity of the condition to be alleviated, or may vary with thesubject's immune response. In general, the goal is to achieve anadequate immune response. For administration by subcutaneous orintradermal infection, the extent of an immune response may bedetermined, for example, by size of delayed local immune skin reactionat the site of injection (e.g., from 0.25 inch to 4 inch diameter). Thedose required to achieve an appropriate immune response may varydepending on the individual (and their immune system) and the responsedesired. Standardized dosages may also be used. In the context ofsubcutaneous or intradermal administration, if the goal is to achieve a2 inch local skin reaction, the total bacterial composition dose may,for example, range from 2 million bacteria (e.g., 0.001 ml of acomposition with a concentration of 2,000 million organisms per ml) tomore than 20,000 million bacteria (e.g., 1 ml of a composition with aconcentration of 20,000 million organisms per ml). The concentrations ofindividual bacterial species or antigens thereof within a compositionmay also be considered. For example, if the concentration of oneparticular pathogenic bacterial species, cell size of that species orantigenic load thereof is much higher relative to the other pathogenicbacterial species in the composition, then the local immune skinreaction of an individual may be likely due to its response to thisspecific bacterial species. In some embodiments, the immune system of anindividual may respond more strongly to one bacterial species within acomposition than another, depending for example on past history ofexposure to infection by a particular species, so the dosage orcomposition may be adjusted accordingly for that individual. However, insome embodiments detailed herein, an immune response will not bemonitored by way of a skin reaction. For example, in some mouse modelsutilized herein, the effective treatment of such animals with antigeniccompositions may not result in corresponding skin reactions. A personskilled in the art will understand that there are alternate ways inwhich an immune response can be monitored beside relying on the presenceor absence of a skin reaction.

For any particular subject, the timing and dose of treatments may beadjusted over time (e.g., timing may be daily, every other day, weekly,monthly) according to the individual need and the professional judgmentof the person administering or supervising the administration of thecompositions. For example, in the context of subcutaneous or intradermaladministration, the compositions may be administered every second day.An initial dose of approximately 0.05 ml may be administeredsubcutaneously, followed by increases from 0.01-0.02 ml every second dayuntil an adequate skin reaction is achieved at the injection site (forexample, a 1 inch to 2 inch diameter delayed reaction of visible rednessat the injection site). Once this adequate immune reaction is achieved,this dosing is continued as a maintenance dose. The maintenance dose maybe adjusted from time to time to achieve the desired visible skinreaction (inflammation) at the injection site. Dosing may be for adosage duration, for example of at least 1 week, 2 weeks, 2 months, 6months, 1, 2, 3, 4, or 5 years or longer.

Oral dosages may for example range from 10 million to 1,000,000 millionorganisms per dose, comprising antigenic determinants of one or morespecies. Oral dosages may be given, for example, from 4 times per day,daily or weekly. Dosing may be for a dosage duration, for example of atleast 1 week, 2 weeks, 2 months, 6 months, 1, 2, 3, 4, or 5 years orlonger.

In some embodiments, the invention may include antigenic compositionsadministered sublingually or by inhalation, or administered to one ormore epithelial tissues (i.e., skin by intradermal or subcutaneousinjection; lung epithelium by inhalation; gastrointestinal mucosa byoral ingestion; mouth mucosa by sublingual administration)simultaneously or sequentially. Accordingly, in some embodiments theantigenic compositions of the invention are administered so as toprovoke an immune response in an epithelial tissue. In some embodiments,one or more epithelial routes of administration may be combined with oneor more additional routes of administration, such as intratumoral,intramuscular or intravenous administration.

In various aspects of the invention, the antigenic compositions that areadministered to a patient may be characterized as having an antigenicsignature, i.e., a combination of antigens or epitopes that aresufficiently specific that the antigenic composition is capable ofeliciting an immune response that is specific to a particular pathogen,such as an adaptive immune response. A surprising and unexpected aspectof the invention is that the non-adaptive or non-specific activation ofthe immune response that is mediated by these specific antigeniccompositions is effective to treat heterologous infections situated inthe tissues in which the particular pathogen is pathogenic.

Routes of administration and dosage ranges set forth herein areexemplary only and do not limit the route of administration and dosageranges that may be selected by medical practitioners. The amount ofactive compound (e.g., pathogenic bacterial species or viruses orantigens thereof) in the composition may vary according to factors suchas the disease state, age, sex, and weight of the individual. Dosageregimens may be adjusted to provide the optimum therapeutic response.For example, a single bolus may be administered, several divided dosesmay be administered over time or the dose may be proportionally reducedor increased as indicated by the exigencies of the therapeuticsituation. It may be advantageous to formulate parenteral compositionsin dosage unit form for ease of administration and uniformity of dosage.

In the case of antigenic formulations (i.e. formulations that provoke animmune response), an immunogenically effective amount of a compound orcomposition of the invention can be provided, alone or in combinationwith other compounds, such as an immunological adjuvant. The compoundmay also be linked with a carrier molecule, such as bovine serum albuminor keyhole limpet hemocyanin to enhance immunogenicity. An antigeniccomposition is a composition that includes materials that elicit adesired immune response. An antigenic composition may select, activateor expand, without limitation: memory B, T cells, neutrophils, monocytesor macrophages of the immune system to, for example, reduce or eliminatethe growth or proliferation of heterologous micro-organisms. In someembodiments, the specific pathogenic microbe, virus, viral antigens,bacteria, bacterial antigens, or compositions thereof of the inventionare capable of eliciting the desired immune response in the absence ofany other agent, and may therefore be considered to be an antigeniccomposition. In some embodiments, an antigenic composition includes asuitable carrier, such as an adjuvant, which is an agent that acts in anon-specific manner to increase the immune response to a specificantigen, or to a group of antigens, enabling the reduction of thequantity of antigen in any given dose, or the reduction of the frequencyof dosage required to generate the desired immune response. A bacterialantigenic composition may include live or dead bacteria capable ofinducing an immune response against antigenic determinants normallyassociated with the bacteria. In some embodiments, an antigeniccomposition may include live bacteria that are of less virulent strains(attenuated), and therefore cause a less severe infection. In someembodiments the antigenic composition may include live, attenuated ordead viruses capable of inducing an immune response against antigenicdeterminants normally associated with the virus.

An antigenic composition comprising killed bacteria for administrationby injection may be made as follows. The bacteria may be grown insuitable media, and washed with physiological salt solution. Thebacteria may then be centrifuged, resuspended in saline solution, andkilled with heat. The suspensions may be standardized by directmicroscopic count, mixed in required amounts, and stored in appropriatecontainers, which may be tested for safety, shelf life, and sterility inan approved manner. In addition to the pathogenic bacterial speciesand/or antigens thereof, a killed bacterial composition suitable foradministration to humans may include 0.4% phenol preservative and/or0.9% sodium chloride. The bacterial composition may also include traceamounts of brain heart infusion (beef), peptones, yeast extract, agar,sheep blood, dextrose, sodium phosphate and/or other media components.

In some embodiments, the bacterial or microbial composition may be usedin tablet or capsule form or drops for oral ingestion, as an aerosol forinhalation, or as drops, aerosol or tablet form for sublingualadministration.

In antigenic compositions comprising bacteria, the concentrations ofspecific bacterial species in compositions for subcutaneous orintradermal injection may be about 1 million to 100,000 millionorganisms per ml, or may be 100 million to 7000 million organisms perml, or may be 500 million to 6000 million organisms per ml, or may be1000 million to 5000 million organisms per ml, or may be 2000 million to4000 million organisms per ml, or any integer within these ranges. Thetotal concentration of bacteria per ml may range from 1 million to100,000 million organisms per ml, or may be 50 million to 7000 millionorganisms per ml, or may be 100 million to 6000 million organisms perml, or may be 500 million to 5000 million organisms per ml, or may be1000 million to 4000 million organisms per ml, or any integer withinthese ranges.

In some embodiments, an antigenic microbial composition for treating aninfection at a particular site (e.g., an infection of the lung tissue)may include pathogenic microbes that commonly, more commonly, or mostcommonly cause infection in that tissue or organ (e.g., infection in thelung tissue i.e., pneumonia).

In general, the pathogenic bacterial species and antigens thereof of theinvention should be used without causing substantial toxicity. Toxicityof the compounds of the invention can be determined using standardtechniques, for example, by testing in cell cultures or experimentalanimals and determining the therapeutic index, i.e., the ratio betweenthe LD50 (the dose lethal to 50% of the population) and the LD100 (thedose lethal to 100% of the population).

As detailed herein and in an aspect of the invention, a method ofcomparing or provoking specific immune responses is provided. The methodinvolves administering to an animal having an organ or tissue amedicament having an antigenic composition, as defined herein. Theantigenic composition may have antigenic determinants selected orformulated so that together the antigenic determinants are specific forat least one microbial pathogen that is pathogenic in the organ ortissue, extracting a quantifiable immune sample from the organ ortissue, measuring a characteristic of the immune response in the organor tissue in the quantifiable immune sample following the administrationof the medicament, and, comparing the characteristic of the immuneresponse in the quantifiable immune sample to a correspondingcharacteristic of the immune response in a reference immune sampleobtained from a corresponding organ or tissue. As used herein, an immunesample would contain sufficient biological material to determine acharacteristic of an immune response. As used herein, a “characteristic”of an immune response can include, without limitation, the particularnumber of a particular immune cell type (e.g., macrophage), or aparticular cellular marker (e.g., upregulation of an integrin) or geneproduct (e.g., a cytokine). The foregoing is provided as an example andis non-limiting.

Optionally, the reference immune sample may be obtained from thecorresponding organ or tissue in the animal prior to the step ofadministering the medicament. In another aspect, the reference immunesample may be obtained from the corresponding organ or tissue in asecond animal such that it is specifically contemplated that at leasttwo animals (i.e., an animal from which a reference immune sample isobtained and a second animal from which a quantifiable immune sample)could be used in the methods described herein. Optionally, the animalmay have an infection situated in the organ or tissue.

Comparing the characteristic of the immune response may involvecomparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells asthese cells are known to those skilled in the art: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Optionally,the macrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages.

Macrophages can be defined as either “M1-like macrophages” or “M2-likemacrophages”. For example, M1-like macrophages are generally understoodby those persons skilled in the art to promote a Th1 CD4+ Tcell-mediated response (see, for e.g., Biswas and Mantovani (2010),Nature Immunology 10:889-96). Moreover, M1-like macrophages aregenerally understood to have efficient antigen presentation capacity,and to be proficient at killing intracellular pathogens (for e.g.,viruses). Moreover, M1-like macrophages are generally understood to beproficient, at least as compared with M2-like macrophages, in playing animmunological role in tumour destruction. Those skilled in the art willappreciate that there are numerous biological markers which can beemployed to differentiate between M1-like macrophages and M2-likemacrophages. For example, and as detailed herein, the expression of Nos2is generally understood to correlate with an M1-like macrophage ascompared with an M2-like macrophage (see, for e.g., Laskin et al. (2010)Annual Rev. Pharmacol. Toxicol. 51: 267-288). Further, and for example,M1-like macrophages are generally understood to produce IL-12 and to beeffectively activated by IFN-γ through the IFN-γR (Biswas and Mantovain,supra).

In contrast to M1-like macrophages, M2-like macrophages promote a Th2CD4+ T cell-mediated response (see, generally: Biswas and Mantovani(2010), Nature Immunology 10:889-96). Moreover, M2-like macrophages aregenerally understood to be effective and encapsulating and clearingextracellular parasites etc. Further, and in comparison to M1-likemacrophages, M2-like macrophages are generally understood by thosepersons skilled in the art as playing a more significant role inimmunoregulation both with respect to T_(reg) and B cells (Biswas andMantovain, supra). Those persons skilled in the art will appreciate thatthere are numerous biological markers which can be employed todifferentiate between M2-like macrophages and M1-like macrophages. Forexample, and as described herein, a diminished expression of Nos2 willgenerally be understood to correlate with M2-like macrophages ascompared with higher expression being generally found in M1-likemacrophages. Further, and as detailed in experiments herein, theexpression of CD206 is generally understood as correlating with M2-likemacrophages (see, for e.g., Choi et al. (2010) Gastroenterology 138(7)2399-409). Further, and as detailed in experiments herein, theexpression of F4/80 is generally understood to correlate with M2-likemacrophages. Further, and for example, M2-like macrophages are generallyunderstood to be effectively activated by IL-4 or by IL-13 throughIL-4Rα (Biswas and Mantovain, supra).

Further, comparing the characteristic of the immune response may involvecomparing a shift in an activation state of macrophages. The shift inthe activation state of macrophages may optionally be characterized as ashift from M2-like macrophages to M1-like macrophages or vice versa.Those persons skilled in the art will appreciate that there are numerousbiological markers that can be employed to monitor the activation ofmacrophages. As detailed herein, those skilled in the art willappreciate that defining a macrophage as being activated towards eithera M1-like phenotype or a M2-like phenotype can be accomplished bychoosing markers that are known to associate with either of therespective phenotypes described herein.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cellular markers on any one or more of the following cells as they arecommonly understood to those persons skilled in the art: inflammatorymonocytes, macrophages, CD11 b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Themacrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages. A person skilled in the art willappreciate that there are numerous cell markers (both extracellular andintracellular) that can be selected which can identify an immuneresponse. For example, as described herein, the marker CD206 isgenerally understood as correlating with M2-like macrophages (see, fore.g., Choi et al. (2010) Gastroenterology 138(7) 2399-409).

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cytokines produced by any one or more of the following cells as they arecommonly understood to those persons skilled in the art: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Those personsskilled in the art will appreciate that cytokines refer to smallcell-signalling protein molecules and that there are numerous cytokinesknown in the art. For example, cytokines have been grouped into type 1and type 2 classifications based on their role in immunologicalresponses. Common type 1 cytokines include IFN-γ and TGF-β. Common type2 cytokines include, but are not limited to IL-4 and IL-13. Cytokinescan be detected by numerous methodologies known to those persons skilledin the art. For example, and as detailed herein, ELISA experiments wereutilized to determine cytokine production from lung tissue.

As detailed herein, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages as has beendefined herein. Optionally, the cytokines are produced as a result of ashift in an activation state of the macrophages. Optionally, themacrophages shift from being M2-like macrophages to being M1-likemacrophages. Further and optionally, the macrophages shift from beingM1-like macrophages to being M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,differential gene expression produced by any one or more of thefollowing cells as they are commonly understood to those persons skilledin the art: inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells,dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells,or NK cells. The macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. The term“differential gene expression” is understood to mean an appreciabledifference between the expression of a particular gene of interest fromat least two experimental conditions. For example, if under a firstexperimental condition a particular gene has a defined expression levelas defined by gene expression methods used by those persons skilled inthe art and if under a second experimental condition the same gene hasan appreciable difference in its expression level, then there isdifferential expression of the gene of interest. Those persons skilledin the art will understand that there are numerous methodologies withwhich to detect differential gene expression. For example, commerciallyavailable quantitative PCR techniques can be used as detailed hereinwith respect to determining the relative Nos2/Arg1 ratios. Optionally,the differential gene expression is produced as a result of a shift inan activation state of the macrophages. Optionally, macrophages mayshift from being M2-like macrophages to being M1-like macrophages asthose terms have been defined herein.

In another embodiment, the medicament may be administered at anadministration site in successive doses given at a dosage interval ofbetween one hour and one month, over a dosage duration of at least oneweek. Optionally, the medicament may be administered intradermally orsubcutaneously. Optionally, the medicament may be administered in a doseso that each dose is effective to cause a visible localized inflammatoryimmune response at the administration site. Optionally, the medicamentmay be administered so that visible localized inflammation at theadministration site occurs within 1 to 48 hours. However, a visiblelocalized inflammatory immune response may not always be present in allcircumstances despite an immune response being initiated. Those skilledin the art will appreciate that there are other methods by which themounting of an immune response can be monitored. For example, theprofile (and relative change in characterization) of immune cells from asubject undergoing an immune reaction can be compared with those from asubject that is not undergoing an immune reaction.

Further and optionally with respect to the methods disclosed herein, theanimal may be a vertebrate, such as a mammal. Optionally, the animal maybe a human or a mouse.

In another aspect, a method of selecting a therapeutic preparationsuitable for treating an individual for an infection in a specific organor tissue is provided. The method involves providing an animal having aninfection situated in a specific organ or tissue, providing a testpreparation having one or more antigenic determinants of a microbialpathogen which is pathogenic in the corresponding specific organ ortissue in a healthy individual, measuring a characteristic of the immuneresponse in a reference immune sample obtained from the organ or tissueof the animal, administering the test preparation to the animal,measuring a characteristic of the immune response in a quantifiableimmune sample obtained from a corresponding organ or tissue of theanimal, comparing the characteristic of the immune response in the inthe reference and quantifiable immune samples, and treating an enhancedcharacteristic of the immune response in the quantifiable immune samplecompared to the reference immune sample as an indication of thesuitability of the test preparation as a therapeutic preparation.Optionally, the animal is sacrificed before the quantifiable immunesample has been obtained.

Optionally, comparing the characteristic of the immune response mayinvolve comparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells asthey are commonly understood to those persons skilled in the art:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages as those termshave been defined herein. Optionally, comparing the characteristic ofthe immune response may involve comparing a shift in an activation stateof macrophages. Optionally, the macrophages may shift from being M2-likemacrophages to being M1-like macrophages. Further and optionally, themacrophages may shift from being M1-like macrophages to being M2-likemacrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cellular markers on any one or more of the following cells as they arecommonly understood to those persons skilled in the art: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Optionally,the macrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages as those terms have been definedherein.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the quantifiable and reference immune samples,cytokines produced by any one or more of the following cells:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages as those terms have beendefined herein. Optionally, the cytokines are produced as a result of ashift in an activate state of the macrophages. Optionally, themacrophages may shift from being M2-like macrophages to being M1-likemacrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, differential gene expression produced by any one or moreof the following cells: inflammatory monocytes, macrophages, CD11b+Gr-1+ cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells,CD8+ T cells, or NK cells. Optionally, the macrophages may include anyone or more of the following: M1-like macrophages or M2-like macrophagesas those terms have been defined herein. Optionally, the differentialgene expression may be produced as a result of a shift in an activationstate of the macrophages. Optionally, the macrophages may shift frombeing M2-like macrophages to being M1-like macrophages. Further andoptionally, the macrophages may shift from being M1-like macrophages tobeing M2-like macrophages.

In another aspect, a method of selectively targeting an immune responseto an infected tissue or an organ in a human subject is provided. Themethod involves administering to the subject a medicament having aneffective amount of a microbial pathogen antigenic composition, whereinthe microbial pathogen may be pathogenic in the specific infected organor tissue of the subject and the antigenic composition comprisesantigenic determinants that together are specific for the microbialpathogen. Optionally, the antigenic composition may include a wholekilled bacterial cell composition. Optionally, the medicament may beadministered to the subject in an amount and for a time that iseffective to up-regulate an immune response in the infected organ ortissue of the subject. Optionally, the method may further involvemeasuring a characteristic of the immune response.

In another aspect, a method for treating a human subject for aninfection situated in a tissue or an organ is provided. The methodinvolves administering to the subject a medicament having an effectiveamount of a microbial pathogen antigenic composition comprising a wholekilled bacterial cell composition, wherein the microbial pathogen ispathogenic in the specific organ or tissue of the subject within whichthe infection is situated. The medicament may be administered to thesubject in an amount and for a time that is effective to modulate animmune response. Optionally, the modulation of the immune response mayinvolve a shift in the activation state of macrophages. Optionally, themodulation of the immune response may involve shifting from a M2-likemacrophage response to a M1-like macrophage response. The modulation ofthe immune responses may involve shifting from a M1-like macrophageresponse to a M2-like macrophage response. Optionally, the method mayfurther involve measuring a characteristic of the immune response.

Optionally, comparing the characteristic of the immune response mayinvolve comparing, in the quantifiable and reference immune samples, anindication of the numbers of any one or more of the following cells asthey are commonly understood to those persons skilled in the art:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages as those termshave been defined herein. Optionally, comparing the characteristic ofthe immune response may involve comparing a shift in an activation stateof macrophages. Further and optionally, the macrophages may shift frombeing M2-like macrophages to being M1-like macrophages. Optionally, themacrophages may shift from being M1-like macrophages to being M2-likemacrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, cellular markers on any one or more of the followingcells as they are commonly understood to those persons skilled in theart: inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages as those terms have beendefined herein. Optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, cytokines produced by any one or more of the followingcells as they are commonly understood to those persons skilled in theart: inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Further,cytokines may be produced as a result of a shift in an activation stateof the macrophages. The macrophages may shift from being M2-likemacrophages to being M1-like macrophages. Optionally, the macrophagesmay shift from being M1-like macrophages to being M2-like macrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the quantifiable and referenceimmune samples, differential gene expression produced by any one or moreof the following cells as they are commonly understood to those personsskilled in the art: inflammatory monocytes, macrophages, CD11b+ Gr-1+cells, dendritic cells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ Tcells, or NK cells. The macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages. Optionally, thedifferential gene expression may be produced as a result of a shift inan activation state of the macrophages. Further and optionally, themacrophages may shift from being M2-like macrophages to being M1-likemacrophages. The macrophages may shift from being M1-like macrophages tobeing M2-like macrophages.

In another aspect, a method of monitoring efficacy of a treatment regimein an individual being treated for an infection in a specific organ ortissue is provided. The method involves measuring a characteristic of animmune response in a post-treatment immune sample obtained from thespecific organ or tissue after the individual has been subject to thetreatment regime for a period of time, wherein the presence of acharacteristic of the immune response which is greater in magnitude thanwould be expected had the individual not been subject to the treatmentregime, is indicative of the efficacy of the treatment regime; and thetreatment regime involves administering a preparation comprising one ormore antigenic determinants of a microbial pathogen which is pathogenicin the corresponding specific organ or tissue in a healthy subject.

The method detailed herein may further involve measuring thecharacteristic of the immune response in a pre-treatment referencesample, wherein the pre-treatment reference sample was obtained from thespecific organ or tissue before, at the same time as or aftercommencement of the treatment regime, but prior to obtaining thepost-treatment immune sample, and comparing the characteristic of theimmune response in the pre-treatment and post-treatment samples, whereinan increase in the magnitude of the immune response in thepost-treatment immune sample compared to the pre-treatment referencesample is indicative of the efficacy of the treatment regime.Optionally, measuring the characteristic of the immune response mayinvolve determining an indication of the number of inflammatorymonocytes in a sample of the organ or tissue. Optionally, measuring thecharacteristic of the immune response may involve determining anindication of the number of macrophages in a sample of the organ ortissue. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages.

Optionally, measuring the characteristic of the immune response mayinvolve determining an indication of the number of CD11b+ Gr-1+ cells ina sample of the organ or tissue or determining an indication of thenumber of dendritic cells in a sample of the organ or tissue. Furtherand optionally, measuring the characteristic of the immune response mayinvolve determining an indication of the number of CD11c+ MHC class II+cells in a sample of the organ or tissue or determining an indication ofthe number of CD4+ T cells in a sample of the organ or tissue ordetermining an indication of the number of CD8+ T cells in a sample ofthe organ or tissue.

Optionally, measuring the magnitude of the immune response may involvedetermining an indication of the number of NK cells in a sample of theorgan or tissue. Further and optionally, comparing the characteristic ofthe immune response may involve identifying, in the reference and immunesamples, cellular markers on any one or more of the following cells asthey are commonly understood to those persons skilled in the art:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. Optionally, the macrophages may include any one or more of thefollowing: M1-like macrophages or M2-like macrophages.

Further and optionally, comparing the characteristic of the immuneresponse may involve identifying, in the reference and immune samples,cytokines produced by any one or more of the following cells as they arecommonly understood to those persons skilled in the art: inflammatorymonocytes, macrophages, CD11b+ Gr-1+ cells, dendritic cells, CD11c+ MHCclass II+ cells, CD4+ T cells, CD8+ T cells, or NK cells. Themacrophages may include any one or more of the following: M1-likemacrophages or M2-like macrophages. Optionally, the cytokines may beproduced as a result of a shift in an activation state of themacrophages. The macrophages may shift from being M2-like macrophages tobeing M1-like macrophages. Further and optionally, the macrophages mayshift from being M1-like macrophages to being M2-like macrophages.

Optionally, comparing the characteristic of the immune response mayinvolve identifying, in the reference and immune samples, differentialgene expression produced by any one or more of the following cells asthey are commonly understood to those persons skilled in the art:inflammatory monocytes, macrophages, CD11b+ Gr-1+ cells, dendriticcells, CD11c+ MHC class II+ cells, CD4+ T cells, CD8+ T cells, or NKcells. The macrophages may include any one or more of the following:M1-like macrophages or M2-like macrophages. The differential geneexpression may be produced as a result of a shift in an activation stateof the macrophages. The macrophages may shift from being M2-likemacrophages to being M1-like macrophages. Optionally, the macrophagesmay shift from being M1-like macrophages to being M2-like macrophages.

The viral pathogen utilized herein may be, without limitation:influenza, adenovirus, respiratory syncytial virus, parainfluenza,monkeypox, herpes simplex virus (1 and 2), Varicella zoster,cytomegalovirus, Epstein-Barr virus, coronavirus, human metapneumovirus,Hendra virus, Nipah virus, Hantavirus, Lassa virus, human T-celllymphotrophic virus, cocksackievirus, echovirus, enterovirus, orrhinovirus, or any virus that is pathogenic in the lung.

The bacterial pathogen utilized herein may be, without limitation:Streptococcus pneumoniae, Moraxella catarrhalis, Mycoplasma pneumoniae,Klebsiella pneumoniae, Haemophilus influenza, Staphylococcus aureus,Chlamydia pneumoniae, Legionella pneumophila, or Bordatella pertussis orany bacterium that is pathogenic in the lung.

The fungal pathogen utilized herein may be, without limitation:Aspergillus fumigatus, Blastomyces sp., Coccidiodes immitis, Coccidiodesposadasii, Cryptococcus neoformans, Cryptococcus gattii, Fusarium sp.,Histoplasma capsulatum, Paecilomyces sp., Paracoccidioides brasiliensis,Penicillium mameffei, Pneumocystis jiroveci, Pseudallescheria boydii,Scedosporium apiospermum, Rhizopus sp., Mucor sp., Absidia sp.,Cunninghamella sp., Scedosporium prolificans, Stachybotrys chartarum,Trichoderma longibrachiatium, Trichosporon sp., or any fungus that ispathogenic in the lung.

In various aspects, embodiments of the invention relate to compositionscomprising components of organisms that may cause infections of thegastrointestinal tract, so that the organism may be characterized as apathogen. However, an organism that is in some cases pathogenic may notalways cause disease. Most animals are colonized to some degree by otherorganisms, such as bacteria, which generally exist in symbiotic orcommensal relationships with the host animal. Thus, many species ofnormally harmless bacteria are found in healthy animals, and are usuallylocalized to the surface of specific organs and tissues. Often, thesebacteria aid in the normal functioning of the body. For example, inhumans, symbiotic Escherichia coli bacteria may be found in theintestine, where they promote immunity and reduce the risk of infectionwith more virulent pathogens.

Bacteria that are generally harmless, such as Escherichia coli, cancause infection in healthy subjects, with results ranging from mild tosevere infection to death. Whether or not an organism, such as abacterium, is pathogenic (i.e., causes infection) depends to some extenton factors such as the route of entry and access to specific host cells,tissues, or organs; the intrinsic virulence of the bacterium; the amountof the bacteria present at the site of potential infection; or thehealth of the host animal. Thus, organisms that are normally harmlesscan become pathogenic given favorable conditions for infection, and evenvirulent organisms may require specific circumstances to causeinfection. Accordingly, organisms that are members of the normal floracan be pathogens when they move beyond their normal ecological role inthe endogenous flora. For example, endogenous species can causeinfection outside of their ecological niche in regions of anatomicalproximity, for example by contiguous spread. When this occurs, and inthe context of the present invention, these normally harmless endogenousorganisms are considered pathogenic.

Specific organisms, such as bacterial species, viruses, worms, andprotozoa are known to cause infections in specific regions of the GIT inotherwise healthy subjects. Examples of organisms that commonly causeinfections in specific regions of the GIT are listed below; it will beunderstood that these examples are not intended to be limiting and thata skilled person would be able to readily recognize and identifyinfectious or pathogenic organisms that cause infections, or commonlycause infections, in various regions of the GIT in healthy adults, basedfor example on knowledge about particular patient populations, asrepresented for example by the following publications: Manual ofClinical Microbiology 8th Edition, Patrick Murray, Ed., 2003, ASM PressAmerican Society for Microbiology, Washington D.C., USA; Mandell,Douglas, and Bennett's Principles and Practice of Infectious Diseases5th Edition, G. L. Mandell, J. E. Bennett, R. Dolin, Eds., 2000,Churchill Livingstone, Philadelphia, Pa., USA, all of which areincorporated by reference herein.

Infections of the mouth are commonly caused by the following bacterialspecies: Prevotella melaninogenicus, anaerobic streptococci, viridansstreptococci, Actinomyces spp., Peptostreptococcus spp., or Bacteroidesspp., or other oral anaerobes; or viral pathogens: herpes simplex,coxsackieviruses, or Epstein-Barr.

Infections of the esophagus are commonly caused by the followingbacterial species: Actinomyces spp., Mycobacterium avium, Mycobacteriumtuberculosis, or Streptococcus spp.; or viral pathogens:cytomegalovirus, herpes simplex, or varicella-zoster.

Infections of the stomach are commonly caused by the following bacterialspecies: Streptococcus pyogenes or Helicobacter pylori; or viralpathogens: cytomegalovirus, herpes simplex, Epstein-Barr, rotaviruses,noroviruses, or adenoviruses.

Infections of the small bowel are commonly caused by the followingbacterial species: Escherichia coli, Clostridium difficile, Bacteroidesfragilis, Bacteroides vulgatus, Bacteroides thetaiotaomicron,Clostridium perfringens, Salmonella enteriditis, Yersiniaenterocolitica, or Shigella flexneri; or viral pathogens: adenoviruses,astroviruses, caliciviruses, noroviruses, rotaviruses, orcytomegalovirus.

Infections of the colon/rectum are commonly caused by the followingbacterial species: Escherichia coli, Clostridium difficile, Bacteroidesfragilis, Bacteroides vulgatus, Bacteroides thetaiotaomicron,Clostridium perfringens, Salmonella enteriditis, Yersiniaenterocolitica, or Shigella flexneri; or viral pathogens: adenoviruses,astroviruses, caliciviruses, noroviruses, rotaviruses, orcytomegalovirus.

Infections of the anus are commonly caused by the following bacterialspecies: Streptococcus pyogenes, Bacteroides spp., Fusobacterium spp.,anaerobic streptococci, Clostridium spp., Escherichia coli, Enterobacterspp., Pseudomonas aeruginosa, or Treponema pallidum; or viral pathogens:herpes simplex.

Organisms such as bacteria are often classified operationally ascollections of similar strains (which generally refer to groups ofpresumed common ancestry with identifiable physiological but usually notmorphological distinctions, and which may be identified usingserological techniques against bacterial surface antigens). Thus, eachbacterial species (e.g., Escherichia coli) has numerous strains (orserotypes), which may differ in their ability to cause infection ordiffer in their ability to cause infection in a particular organ/site.Certain strains of Escherichia coli are more likely to causegastrointestinal infection/diarrhea, including enterotoxigenic E. coli(ETEC), enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli(EHEC), Shiga toxin-producing E. coli (STEC), enteroaggregative E. coli(EAEC), enteroinvasive E. coli (EIEC) and diffuse adhering E. coli(DAEC). In accordance with the present invention, one or more of anETEC, EPEC, EHEC, STEC, EAEC, EIEC or DAEC strains of E. coli (i.e.,strains that cause colon infection), may be chosen for a formulation totreat and a heterologous microbial infection, such as an infection ofthe GIT, for example an IBD-related microbial infection. For example, anon-ETEC strain of E. coli may be used to prepare an antigenicformulation for treating an infection by an ETEC strain of E. coli.Similarly, non-EPEC, non-EHEC, non-STEC, non-EAEC, non-EIEC or non-DAECstrains of E. coli may be used to formulate an antigenic formulation fortreating an infection by, respectively, an EPEC, EHEC, STEC, EAEC, EIECor DAEC strain of E. coli.

Similarly, there may be numerous subtypes of specific viruses, worms, orprotozoa, which are associated with disease in a particular population,and are therefore amenable for use in the present invention.

The compositions of the invention include antigens of organisms that arepathogenic in a specific region of the body, such as the GIT. Thecompositions may include the components of whole organisms, whole cellsor whole virions, or may include extracts or preparations of theorganisms, such as cell wall or cell membrane extracts, or exotoxins.The compositions may also include one or more isolated antigens fromthese organisms. Pathogenic organisms may be available commercially (forexample from the American Type Culture Collection, Manassas, Va., USA),or may be clinical isolates from subjects having an infection.

The compositions of the invention derived from pathogens can be providedalone or in combination with other compounds (for example, nucleic acidmolecules, small molecules, peptides, or peptide analogues), in thepresence of a liposome, an adjuvant, or any pharmaceutically acceptablecarrier, in a form suitable for administration to mammals, for example,humans. As used herein “pharmaceutically acceptable carrier” or“excipient” includes any and all solvents, dispersion media, coatings,antibacterial and antifungal agents, isotonic and absorption delayingagents, and the like that are physiologically compatible. The carriercan be suitable for any appropriate form of administration, includingsubcutaneous, intradermal, intravenous, parenteral, intraperitoneal,intramuscular, sublingual, inhalational, or oral administration.Pharmaceutically acceptable carriers include sterile aqueous solutionsor dispersions and sterile powders for the extemporaneous preparation ofsterile injectable solutions or dispersion. The use of such media andagents for pharmaceutically active substances is well known in the art.Except insofar as any conventional media or agent is incompatible withthe active compound (i.e., the specific bacteria, bacterial antigens, orcompositions thereof of the invention), use thereof in thepharmaceutical compositions of the invention is contemplated.Supplementary active compounds can also be incorporated into thecompositions.

Methods well known in the art for making formulations are found in, forexample, “Remington's Pharmaceutical Sciences” (20th edition), ed. A.Gennaro, 2000, Mack Publishing Company, Easton, Pa. Formulations forparenteral administration may, for example, contain excipients, sterilewater, or saline, polyalkylene glycols such as polyethylene glycol, oilsof vegetable origin, or hydrogenated napthalenes. Biocompatible,biodegradable lactide polymer, lactide/glycolide copolymer, orpolyoxyethylene-polyoxypropylene copolymers may be used to control therelease of the compounds. Other potentially useful parenteral deliverysystems for include ethylene-vinyl acetate copolymer particles, osmoticpumps, implantable infusion systems, and liposomes. Formulations forinhalation may contain excipients, for example, lactose, or may beaqueous solutions containing, for example, polyoxyethylene-9-laurylether, glycocholate and deoxycholate, or may be oily solutions foradministration in the form of nasal drops, or as a gel. For therapeuticor prophylactic compositions, the formulations may be administered to anindividual in an amount effective to prevent, stop or slow progressionof a microbial infection.

An “effective amount” of a pathogenic species or antigen thereofaccording to the invention includes a therapeutically effective amountor a prophylactically effective amount. A “therapeutically effectiveamount” refers to an amount effective, at dosages and for periods oftime necessary, to achieve the desired therapeutic result, such asreduction or elimination of symptoms of a microbial infection. Atherapeutically effective amount of a pathogenic species or antigen(s)thereof may vary according to factors such as the disease state, age,sex, and weight of the individual, and the ability of the compound toelicit a desired response in the individual. Dosage regimens may beadjusted to provide the optimum therapeutic response. A therapeuticallyeffective amount may also be one in which any toxic or detrimentaleffects of the pathogenic species or antigen thereof are outweighed bythe therapeutically beneficial effects. A “prophylactically effectiveamount” refers to an amount effective, at dosages and for periods oftime necessary, to achieve the desired prophylactic result, such asprevention of a microbial infection. Typically, a prophylactic dose isused in subjects prior to or at an earlier stage of a microbialinfection, so that a prophylactically effective amount may be less thana therapeutically effective amount.

For administration by subcutaneous or intradermal injection, anexemplary range for therapeutically or prophylactically effectiveamounts of one or more pathogenic bacterial species may be about 1million to 100,000 million organisms per ml, or may be 100 million to7000 million organisms per ml, or may be 500 million to 6000 millionorganisms per ml, or may be 1000 million to 5000 million organisms perml, or may be 2000 million to 4000 million organisms per ml, or anyinteger within these ranges. The total concentration of bacteria per mlmay range from 1 million to 100,000 million organisms per ml, or may be50 million to 7000 million organisms per ml, or may be 100 million to6000 million organisms per ml, or may be 500 million to 5000 millionorganisms per ml, or may be 1000 million to 4000 million organisms perml, or any integer within these ranges. The range for therapeutically orprophylactically effective amounts of antigens of a pathogenic bacterialspecies may be any integer from 0.1 nM-0.1 M, 0.1 nM-0.05M, 0.05 nM-15μM or 0.01 nM-10 μM.

It is to be noted that dosage concentrations and ranges may vary withthe severity of the condition to be alleviated, or may vary with thesubject's immune response. In general, the goal is to achieve anadequate immune response. For administration by subcutaneous orintradermal infection, the extent of an immune response may bedetermined, for example, by size of delayed local immune skin reactionat the site of injection (e.g., from 0.25 inch to 4 inch diameter). Thedose required to achieve an appropriate immune response may varydepending on the individual (and their immune system) and the responsedesired. Standardized dosages may also be used.

In the context of subcutaneous or intradermal administration, if thegoal is to achieve a 2 inch local skin reaction, using a bacterialcomposition, the total dose may, for example, range from 2 millionbacteria (e.g., 0.001 ml of a composition with a concentration of 2,000million organisms per ml) to more than 20,000 million bacteria (e.g., 1ml of an antigenic composition with a concentration of 20,000 millionorganisms per ml). The concentrations of individual bacterial species orantigens thereof within a composition may also be considered. Forexample, if the concentration of one particular pathogenic bacterialspecies, cell size of that species or antigenic load thereof is muchhigher relative to the other pathogenic bacterial species in theantigenic composition, then the local immune skin reaction of anindividual may be likely due to its response to this specific bacterialspecies. In some embodiments, the immune system of an individual mayrespond more strongly to one bacterial species within a composition thananother, depending for example on past history of exposure to infectionby a particular species, so the dosage or composition may be adjustedaccordingly for that individual.

For any particular subject, the timing and dose of treatments may beadjusted over time (e.g., timing may be daily, every other day, weekly,monthly) according to the individual need and the professional judgementof the person administering or supervising the administration of thecompositions. For example, in the context of subcutaneous or intradermaladministration, the compositions may be administered every second day.An initial dose of approximately 0.05 ml may be administeredsubcutaneously, followed by increases from 0.01-0.02 ml every second dayuntil an adequate skin reaction is achieved at the injection site (forexample, a 1 inch to 2 inch diameter delayed reaction of visible rednessat the injection site). Once this adequate immune reaction is achieved,this dosing is continued as a maintenance dose. The maintenance dose maybe adjusted from time to time to achieve the desired visible skinreaction (inflammation) at the injection site. Dosing may be for adosage duration, for example of at least 2 weeks, 2 months, 6 months, 1,2, 3, 4, or 5 years or longer.

In some embodiments, the invention may include antigenic compositionsadministered to one or more epithelial tissues by a non-enteric route.For example: to skin by intradermal or subcutaneous injection; to lungepithelium by inhalation. Accordingly, in some embodiments the antigeniccompositions of the invention are administered so as to provoke animmune response in a non-enteric tissue, such as an epithelial tissue.In some embodiments, one or more non-enteric routes of administrationmay be combined with one or more additional routes of administration,such as intramuscular or intravenous administration.

In various aspects of the invention, the antigenic compositions that areadministered to a patient may be characterized as having an antigenicsignature, i.e., a combination of antigens or epitopes that issufficiently specific that the antigenic composition is capable ofeliciting an immune response that is specific to a particular pathogen,such as an adaptive immune response.

The amount of active compound (e.g., bacterial species, viruses,protozoa or helminths, or antigens thereof) in compositions of theinvention may vary according to factors such as the disease state, age,sex, and weight of the individual. Dosage regimens may be adjusted toprovide the optimum therapeutic response. For example, a single bolusmay be administered, several divided doses may be administered over timeor the dose may be proportionally reduced or increased as indicated bythe exigencies of the therapeutic situation. It may be advantageous toformulate parenteral compositions in dosage unit form for ease ofadministration and uniformity of dosage.

In the case of antigenic formulations, an immunogenically effectiveamount of a compound of the invention can be provided, alone or incombination with other compounds, with an immunological adjuvant. Thecompound may also be linked with a carrier molecule, such as bovineserum albumin or keyhole limpet hemocyanin to enhance immunogenicity. Anantigenic composition is a composition that includes materials thatelicit a desired immune response. An antigenic composition may select,activate or expand memory B, T cells, neutrophils, monocytes ormacrophages of the immune system to, for example, reduce or eliminatethe symptoms of the microbial infection. In some embodiments, thespecific pathogenic microbe, virus, viral antigens, bacteria, bacterialantigens, or compositions thereof of the invention are capable ofeliciting the desired immune response in the absence of any other agent,and may therefore be considered to be an antigenic composition. In someembodiments, an antigenic composition includes a suitable carrier, suchas an adjuvant, which is an agent that acts in a non-specific manner toincrease the immune response to a specific antigen, or to a group ofantigens, enabling the reduction of the quantity of antigen in any givendose, or the reduction of the frequency of dosage required to generatethe desired immune response. A bacterial antigenic composition mayinclude live or dead bacteria capable of inducing an immune responseagainst antigenic determinants normally associated with the bacteria. Insome embodiments, an antigenic composition may include live bacteriathat are of less virulent strains (attenuated), and therefore cause aless severe infection. In some embodiments the antigenic composition mayinclude live, attenuated or dead viruses capable of inducing an immuneresponse against antigenic determinants normally associated with thevirus.

An antigenic composition comprising killed organisms for administrationby injection may be made as follows. The organism may be grown insuitable media, and washed with physiological salt solution. Theorganism may then be centrifuged, resuspended in saline solution, andkilled with heat. The suspensions may be standardized by directmicroscopic count, mixed in required amounts, and stored in appropriatecontainers, which may be tested for safety, shelf life, and sterility inan approved manner. In addition to the organism and/or antigens thereof,a killed preparation suitable for administration to humans may includephenol preservative (for example 0.4%) and/or sodium chloride (forexample on the order of 0.9%). The composition may also for exampleinclude trace amounts of brain heart infusion (beef), peptones, yeastextract, agar, sheep blood, dextrose, sodium phosphate and/or othermedia components.

In some embodiments, the antigenic composition may be used as an aerosolfor inhalation.

In general, the compositions of the invention should be used withoutcausing substantial toxicity. Toxicity of the compounds of the inventioncan be determined using standard techniques, for example, by testing incell cultures or experimental animals and determining the therapeuticindex, i.e., the ratio between the LD50 (the dose lethal to 50% of thepopulation) and the LD100 (the dose lethal to 100% of the population).

In some embodiments, bacteria that are members of the endogenous floraof a particular region of the GIT may be used to formulate antigeniccompositions of the invention. The rows of Table 1 list a number ofbacterial species, together with the biological regions in which eachspecies may form a part of the endogenous flora. For example,Abiotrophia spp. are typically members of the endogenous flora of themouth.

TABLE 1 Human Bacterial Normal Flora (Endogenous Bacterial HumanPathogens) Duodenum/ Bacterial species Mouth Stomach Jejunum Ileum ColonCFU/mL 10{circumflex over ( )}5 10{circumflex over ( )}2 10{circumflexover ( )}5 10{circumflex over ( )}8 10{circumflex over ( )}11Abiotrophia spp + Acholeplasma + laidlawii Acidaminococcus + + + +fermentans Acinetobacter + + + + spp. Actinobacillus + spp.Actinobaculum + + + + spp. Actinomyces + + + + spp. Aeromonas spp. + + +Anaerorhabdus + + furcosus Anaerococcus + + hydrogenalisAnaerococcus + + lactolyticus Anaerococcus + + prevotii Atopobiumspp. + + + + Bacillus spp. + + Bacteroides + + caccae Bacteroides + +distasonis Bacteroides + + eggerthii Bacteroides + + fragilisBacteroides + + merdae Bacteroides + + ovatus Bacteroides + +splanchnicus Bacteroides + + thetaiotaomicron Bacteroides + + vulgatusBifidobacterium + + + adolescentis Bifidobacterium + + + bifidumBifidobacterium + + + breve Bifidobacterium + + + catenulatumBifidobacterium + + + + dentium Bifidobacterium + + + longumBilophila + + + + wadsworthia Burkholderia + + + cepaciaButyrivibrio + + + fibrisolvens Campylobacter + + + concisusCampylobacter + + + curvus Campylobacter + + + gracilisCampylobacter + + + jejuni Campylobacter + + + rectusCampylobacter + + + + showae Campylobacter + sputorum Capnocytophaga +granulosum Capnocytophaga + gingivalis Campylobacter + haemolyticaCapnocytophaga + + + + ochracea Capnocytophaga + sputigenaCardiobacterium + hominis Cedecea spp + Centipeda + periodontiiCitrobacter + + + freundii Citrobacter + + + koseri Clostridiumspp. + + + Corynebacterium + accolens Corynebacterium + afermentansDesulfomonas + + + pigra Dysgonomonas + + + spp. Eikenella + + + +corrodens Enterobacter + + + aerogenes Enterobacter + + + cloacaeEnterobacter + + + gergoviae Enterobacter + + + sakazakiiEnterobacter + + + taylorae Enterococcus + + + spp. Escherichiacoli + + + Escherichia + + + fergusonii Escherichia + + + hermanniiEscherichia + + + vulneris Eubacterium + + + + spp. Ewingella +americana Finegoldia + + + magnus Fusobacterium + alocisFusobacterium + + + gonidiaformans Fusobacterium + + + mortiferumFusobacterium + + + naviforme Fusobacterium + + + + necrophorumFusobacterium + + nucleatum Fusobacterium + sulci Fusobacterium + + +russii Fusobacterium + + + varium Gardnerella + + + vaginalis Gemella +haemolysans Gemella + + + + morbillorum Globicatella spp. + +Granulicatella + spp. Haemophilus + spp. Hafnia alvei + + + Helcococcuskunzii Helicobacter spp. + + + Kingella spp. + Klebsiella spp. + + + +Lactobacillus + + + + + acidophilus Lactobacillus + breveLactobacillus + casei Lactobacillus + + + + + fermentumLactobacillus + + + + reuteri Lactobacillus + + + + + salivariusLeclercia + + + adecarboxylata Leminorella spp. + + + Leptotrichia +buccalis Megasphaera + + + elsdenii Micrococcus + luteus Micrococcus +lylae Micromonas + micros Mitsuokella + + + multiacidus Mobiluncus + + +curisii Mobiluncus + + + mulieris Moellerella + + + wisconsensisMoraxella + catarrhalis other Moraxella + spp. Morganella + + + morganiiMycoplasma + buccale Mycoplasma + fermentans Mycoplasma + hominisMycoplasma + lipophilum Mycoplasma + orale Mycoplasma + pneumoniaeMycoplasma + salivarium Pantoea + + + agglomerans Pasteurella +multocida Pediococcus + + spp. Peptoniphilus + + + asaccharolyticusPeptostreptococcus + + + + anaerobus Peptostreptococcus + + + productusPorphyromonas + + + + asaccharolytica Porphyromonas + + catoniaePorphyromonas + + endodontalis Porphyromonas + + gingivalisPrevotella + + buccae Prevotella + + buccalis Prevotella + + corporisPrevotella + + dentalis Prevotella + + denticola Prevotella + + enoecaPrevotella + + heparinolytica Prevotella + + intermedia Prevotella + +loescheii Prevotella + + melaninogenica Prevotella + + nigrescensPrevotella oralis + + Prevotella oris + + Prevotella + + oulorumPrevotella + + tannerae Prevotella + + veroralis Prevotella + +zoogleoformans Propionibacterium + propionicum Proteus mirabilis + +Proteus penneri + + Proteus vulgaris + + Providencia + + rettgeriProvidencia + + + stuartii Pseudomonas + + + aeruginosaRetortamonas + + + intestinalis Rothia + dentocariosa Rothia +mucilaginosa Ruminococcus + + + productus Selenomonas + spp.Serratia + + liquefaciens Serratia + + marcescens Serratia odorifera + +Staphylococcus + aureus Staphylococcus + epidermidis Streptococcus + + +agalactiae Streptococcus + + + + anginosus Streptococcus + + + bovisStreptococcus + + + + constellatus Streptococcus + cricetiStreptococcus + crista Streptococcus + equisimilis Streptococcus +gordonii Streptococcus + + + intermedius Streptococcus + + mitisStreptococcus + mutans Streptococcus + oralis Streptococcus +parasanguis Streptococcus + + pyogenes Streptococcus + + salivariusStreptococcus + + sanguis Streptococcus + sobrinus Streptococcus +vestibularis Group C + G + + Streptococci Succinivibrio + + +dextrinosolvens Sutterella spp. + + + Suttonella + indologenesTissierella + + + praeacuta Treponema + denticola Treponema +maltophilum Treponema + socranskii Treponema + vincentii Ureaplasma +urealyticum Veillonella spp. + + + +

Endogenous microbial flora, such as bacteria, have access to tissues forpathogenesis either through contiguous spread or bacteremic spread.Under favorable conditions, all endogenous organisms can becomepathogenic and invade locally and spread by contiguous spread toadjacent tissues and organs. Endogenous bacterial flora of the skin,mouth and colon are the species that are understood to also be amenableto bacteremic spread. Bacteria that are members of a particularendogenous flora domain may therefore cause infection in tissues ororgans to which these bacteria may spread. Accordingly, one aspect ofthe invention involves the use of endogenous microbial pathogens totreat a microbial infection having symptoms localized to a region of theGIT in which the endogenous bacteria may spread to cause infection. Thecolumns of Table 2 list domains for endogenous flora. The rows of Table2 list regions of the GIT within which a microbial infection may besituated. Accordingly, one aspect of the invention involves the use ofendogenous microbial pathogens to formulate antigenic compositions, orthe selection of existing formulations having the pathogens, fortreating a microbial infection situated in the region of the GIT towhich the pathogen may spread to cause an infection. Accordingly, inalternative embodiments, a microbial infection that is symptomatic inthe region listed in the first column of Table 2 may be treated withantigenic compositions comprising antigenic determinants that arespecific for microbial pathogens that are members of the endogenousflora of one or more of the endogenous flora domains listed in the firstrow of Table 2 and indicated with an X or a check mark in theappropriate row.

TABLE 2 Tissue/Organ Pathogenicity of Endogenous Flora Tissue/Duo-denum/ organ site Mouth Stomach Jejunum Ileum Colon Oral x Tonsil xNasophar- ynx/Sinus x Esophagus x Stomach x Small bowel x x Colon/ xRectum Anus x

In accordance with the combined information in Tables 1 and 2, amicrobial infection manifest in a particular region of the GIT set outin column 1 of Table 2 may be treated with antigenic compositionscomprising antigenic determinants of the corresponding bacterial speciesof Table 1, so that the column headings in Table 2 are in effectreplaced with the bacterial species of Table 1.

In some embodiments, pathogens for use in the invention may be exogenousbacterial pathogens. For example, the organisms listed in Table 3 may beused as microbial pathogens to formulate antigenic compositions, orantigenic compositions having those pathogens may selected, for use totreat a microbial infection situated in the region of the GIT listedwith the relevant organism in Table 3. In some embodiments, antigenicdeterminants of both endogenous and exogenous bacterial species targetedto a specific tissue or organ may be used in combination. For example,an antigenic composition derived from, or specific for, Clostridiumdifficile, may be used to treat a microbial infection situated in thecolon.

TABLE 3 Exogenous Bacterial Human Pathogens, and their Sites ofInfection in the GIT Bacterial Species Region of the GIT Aerobacter spp.small bowel, colon, Bacillus anthracis oral, small bowel, colon,hematological Bacillus cereus colon, other Bacillus spp. colon, stomach,small bowel Brucella spp. small bowel, colon Campylobacter coli smallbowel, colon Campylobacter colon jejuni Campylobacter small bowel, colonsputorum Clostridium small bowel, colon, stomach bifermentansClostridium colon, small bowel botulinum Clostridium difficile colonClostridium indolis small bowel, colon, stomach, Clostridium smallbowel, colon, stomach mangenolii Clostridium small bowel, colon, stomachperfringens Clostridium sordellii small bowel, colon, stomachClostridium small bowel, colon, stomach sporogenes Clostridium smallbowel, colon, stomach subterminale Edwarsiella tarda small bowel, colonFrancisella small bowel tularensis Helicobacter pylori stomachLeptospirosis spp. oral Listeria small bowel, colon monocytogenesMycobacterium colon, small bowel bovis Mycobacterium small bowel, colontuberculosis Pediococcus spp. colon Plesiomonas small bowel, colonshigelloides Rickettsia small bowel rickettsiae Salmonella spp. stomach,small bowel, colon Shigella boydii colon Shigella colon dysenteriaeShigella flexneri colon Shigella sonnei colon other Spirillum spp. colonStreptococcus small bowel zooepidemicus Treponema oral, anus pallidumTropheryma small bowel, colon whipplei Vibrio cholerae colon, smallbowel Vibrio fluvialis small bowel, colon Vibrio furnissii small bowel,colon Vibrio hollisae small bowel, colon Vibrio colon, small bowelparahaemolyticus Yersinia small bowel, colon enterocolitica Yersiniasmall bowel, colon pseudotuberculosis

In some embodiments, pathogens for use in the invention may be viralpathogens. Table 4 provides an exemplary list of viral pathogenstogether with the tissue and organ sites for which each viral species isreportedly a pathogen. Accordingly, one aspect of the invention involvesutilizing immunogenic compositions that are specific for the namedviruses to treat a pathology associated with a heterologous infectionsituated in the region of the GIT that is identified adjacent to thename of the virus in Table 4.

TABLE 4 Viral Human Pathogens and Their Sites of Infection Virus Regionof the GIT Herpes Simplex rectum, anus virus (1 and 2) Cytomegalovirussmall bowel, colon/rectum Epstein-Barr virus oral Adenovirus oral, smallbowel, colon Human anus, oral papillomavirus Orthoreoviruses smallbowel, colon, oral Coltiviruses oral Rotaviruses small bowel, colonAlphaviruses small bowel, colon, Coronaviruses oral, small bowel, colonToroviruses small bowel, colon Parainfluenza oral viruses Respiratorysyncytial oral virus Human oral, small bowel, colon metapneumovirusVesicular stomatitis oral, small bowel, colon virus Rabies virus oralInfluenza virus oral Hantaviruses oral Machupo virus small bowel, colonJunin virus small bowel, colon Poliovirus small bowel, colonCoxsackieviruses small bowel, colon Echoviruses oral, small bowel, colonHepatitis A virus small bowel, colon Rhinoviruses oral Noroviruses andsmall bowel, colon other Caliciviruses Astroviruses small bowel, colonPicobirnaviruses small bowel, colon Hepatitis E virus small bowel, colon

The cumulative information in Tables 1 through 4 provides an extensiveidentification of pathogens that may be used in the formulation ofantigenic compositions of the invention, together with an identificationof the region of the GIT in which these organisms are pathogenic, andaccordingly identifies the region of the GIT in which an infection issituated that may be treated with an antigenic anti-microbialformulation of the invention. Pathogens may be selected from endogenouspathogens or exogenous pathogens.

In some embodiments, the pathogen selected for use in antigeniccompositions of the invention may be one that is a common cause of acuteinfection in the region of the GIT in which the microbial infection tobe treated is situated. Table 5 identifies bacterial and viral pathogensof this kind, together with the region of the GIT in which they commonlycause infection. Accordingly, in selected embodiments, a microbialinfection, such as an IBD-related infection, residing in a region of theGIT identified in the first column of Table 5 may be treated with anantigenic composition that comprises antigenic determinants for one ormore of the pathogenic organisms listed in the second column of Table 5.

TABLE 5 Common causes of acute infection (bacteria and viruses) forselected regions of the GIT Selected regions of the GIT Common Bacterialor Viral Pathogens Oral Prevotella melaninogenicus, anaerobicstreptococci, viridans streptococci, Actinomyces spp.,Peptostreptococcus spp., Bacteroides spp., and other oral anaerobesherpes simplex, coxsackieviruses, Epstein-Barr Stomach Streptococcuspyogenes, Helicobacter pylori cytomegalovirus, herpes simplex,Epstein-Barr, rotaviruses, noroviruses, adenoviruses Small Escherichiacoli, Clostridium difficile, Bacteroides fragilis, bowel Bacteroidesvulgatus, Bacteroides thetaiotaomicron, Clostridium perfringens,Salmonella enteriditis, Yersinia enterocolitica, Shigella flexneriadenoviruses, astroviruses, caliciviruses, noroviruses, rotaviruses,cytomegalovirus Colon/ Escherichia coli, Clostridium difficile,Bacteroides fragilis, Rectum Bacteroides vulgatus, Bacteroidesthetaiotaomicron, Clostridium perfringens, Salmonella enteriditis,Yersinia enterocolitica, Shigella flexneri adenoviruses, astroviruses,caliciviruses, noroviruses, rotaviruses, cytomegalovirus AnusStreptococcus pyogenes, Bacteroides spp., Fusobacterium spp., anaerobicstreptococci, Clostridium spp., E. coli, Enterobacter spp., Pseudomonasaeruginosa, Treponema pallidum herpes simplex

The specific organisms which commonly cause infection in a specificregion of the GIT may vary by geographical location. Table 5 is thus notan exhaustive list of common pathogens for all geographic locations andpopulation groups. It is understood that a clinical microbiologistskilled in the art could determine the common pathogenic species in aparticular geographic area or population group for a specific region ofthe GIT in accordance with the invention.

Humans are hosts to a wide range of gastrointestinal parasites,including various protozoa and helminths, which for purposes of thepresent invention constitute pathogens of the GIT (Schafer, T. W.,Skopic, A. Parasites of the small intestine. Curr Gastroenterol Reports2006; 8:312-20; Jernigan, J., Guerrant, R. L., Pearson, R. D. Parasiticinfections of the small intestine. Gut 1994; 35:289-93; Sleisenger &Fordtran's Gastrointestinal and liver disease. 8th ed. 2006; Garcia, L.S. Diagnostic medical parasitology. 5th ed. 2007). Compositions of theinvention may accordingly include antigenic components of variousprotozoa, including for example: Giardia lamblia, Cryptosporidiumparvum, Cryptosporidium hominus, Isospora belli, Sarcocystis species,Coccidian like bodies (Cyclospora species), Enterocytozoon bieneusi,Entamoeba histolytica, Entamoeba dispar, Entamoeba coli, Entamoebahartmanni, Endolimax nana, Iodamoeba bütschlii, Dientameoba fragilis,Blastocystis hominus, Cyclospora cayetanensis, Microsporidia,Trypanosoma cruzi, Chilomastix mesnili, Pentatrichomonas hominis,Balantidium coli. Similarly, compositions of the invention may includeantigenic components of various helminths, including for example:Cestodes (tapeworms), Taenia saginata, Taenia solium, Diphyllobothriumspecies, Hymenolepis nana, Hymenolepis diminuta, Dipylidium caninum,Nematodes (round worms), Ascaris lumbricoides, Strongyloidesstercoralis, Necator americanus, Ancylostoma duodenale, Ancylostomacaninum, Tichuris trichiura, Capillaria philippinensis, Trichostrongylusspecies, Trichinella species, Necator americanus, Anisakis and relatedspecies, Angiostrongylus costaricensis, Enterobius vermicularis,Trematodes (flukes), Fasciolopsis buski, Heterophyes speicies,Echinostoma species, Clonorchis sinensis, Opisthorchis species, Fasciolaspecies, Metagonimus yokogawi, Schistosoma mansoni, Schistosomajaponicum, Schistosoma mekongi, Schistosoma intercalatum, Echinostomaspecies and Paragonimus species.

In accordance with the foregoing, in various aspects, the invention mayinvolve the treatment of a microbial infection, such as a microbialinfection of the GIT, or an IBD-related microbial infection, withformulations of a pathogen is selected from the group consisting of:Acidaminococcus fermentans; Acinetobacter spp.; Actinobaculum spp.;Actinomyces spp.; Aeromonas spp.; Anaerorhabdus furcosus; Anaerococcushydrogenalis; Anaerococcus lactolyticus; Anaerococcus prevotii;Atopobium spp.; Bacillus spp.; Bacteroides caccae; Bacteroidesdistasonis; Bacteroides eggerthii; Bacteroides fragilis; Bacteroidesmerdae; Bacteroides ovatus; Bacteroides splanchnicus; Bacteroidesthetaiotaomicron; Bacteroides vulgatus; Bifidobacterium adolescentis;Bifidobacterium bifidum; Bifidobacterium breve; Bifidobacteriumcatenulatum; Bifidobacterium dentium; Bifidobacterium longum; Bilophilawadsworthia; Burkholderia cepacia; Butyrivibrio fibrisolvens;Campylobacter concisus; Campylobacter curvus; Campylobacter gracilis;Campylobacter jejuni; Campylobacter rectus; Campylobacter showae;Capnocytophaga ochracea; Cedecea spp; Citrobacter freundii; Citrobacterkoseri; Clostridium spp.; Desulfomonas pigra; Dysgonomonas spp.;Eikenella corrodens; Enterobacter aerogenes; Enterobacter cloacae;Enterobacter gergoviae; Enterobacter sakazakii; Enterobacter taylorae;Enterococcus spp.; Escherichia coli; Escherichia fergusonii; Escherichiahermannii; Escherichia vulneris; Eubacterium spp.; Finegoldia magnus;Fusobacterium gonidiaformans; Fusobacterium mortiferum; Fusobacteriumnaviforme; Fusobacterium necrophorum; Fusobacterium nucleatum;Fusobacterium russii; Fusobacterium varium; Gardnerella vaginalis;Gemella morbillorum; Globicatella spp.; Hafnia alvei; Helicobacter spp.;Klebsiella spp.; Lactobacillus acidophilus; Lactobacillus fermentum;Lactobacillus reuteri; Lactobacillus salivarius; Leclerciaadecarboxylata; Leminorella spp.; Megasphaera elsdenii; Mitsuokellamultiacidus; Mobiluncus curisii; Mobiluncus mulieris; Moellerellawisconsensis; Morganella morganii; Pantoea agglomerans; Pediococcusspp.; Peptoniphilus asaccharolyticus; Peptostreptococcus anaerobus;Peptostreptococcus productus; Porphyromonas asaccharolytica; Proteusmirabilis; Proteus penneri; Proteus vulgaris; Providencia rettgeri;Providencia stuartii; Pseudomonas aeruginosa; Retortamonas intestinalis;Ruminococcus productus; Serratia liquefaciens; Serratia marcescens;Serratia odorifera; Streptococcus agalactiae; Streptococcus anginosus;Streptococcus bovis; Streptococcus constellatus; Streptococcusintermedius; Group C+G Streptococci; Succinivibrio dextrinosolvens;Sutterella spp.; Tissierella praeacuta; Veillonella spp.; Aerobacterspp.; Bacillus anthracis; Bacillus cereus; other Bacillus spp.; Borreliarecurrentis; Brucella spp.; Campylobacter coli; Campylobacter fetus;Campylobacter jejuni; Campylobacter sputorum; Clostridium bifermentans;Clostridium botulinum; Clostridium difficile; Clostridium indolis;Clostridium mangenolii; Clostridium perfringens; Clostridium sordellii;Clostridium sporogenes; Clostridium subterminale; Edwarsiella tarda;Francisella tularensis; Listeria monocytogenes; Mycobacterium bovis;Mycobacterium tuberculosis; Pediococcus spp.; Plesiomonas shigelloides;Rickettsia rickettsiae; Salmonella spp.; Shigella boydii; Shigelladysenteriae; Shigella flexneri; Shigella sonnei; other Spirillum spp.;Streptococcus zooepidemicus; Tropheryma whipplei; Vibrio cholerae;Vibrio fluvialis; Vibrio furnissii; Vibrio hollisae; Vibrioparahaemolyticus; Yersinia enterocolitica; Yersinia pseudotuberculosis;Herpes Simplex virus (1 and 2); Cytomegalovirus; Adenovirus;Orthoreoviruses; Rotaviruses; Alphaviruses; Coronaviruses; Toroviruses;Human metapneumovirus; Vesicular stomatitis virus; Machupo virus; Juninvirus; Poliovirus; Coxsackieviruses; Echoviruses; Hepatitis A virus;Noroviruses and other Caliciviruses; Astroviruses; Picobirnaviruses; andHepatitis E virus.

In alternative aspects, the invention may involve the treatment of amicrobial infection, such as an infection of the GIT, for example anIBD-related infection, with formulations wherein the pathogen isselected from the group of common small and larger bowel pathogens, forexample the group consisting of: Escherichia coli, Clostridiumdifficile, Bacteroides fragilis, Bacteroides vulgatus, Bacteroidesthetaiotaomicron, Clostridium perfringens, Salmonella enteriditis,Yersinia enterocolitica, Shigella flexneri; adenoviruses, astroviruses,caliciviruses, noroviruses, rotaviruses, and cytomegalovirus.

In selected embodiments, the invention involves diagnostic steps toassess a patient's previous exposure to an organism. For example, thediagnostic steps may include taking a medical history of exposure toselected pathogens, and/or evaluating a patient's immune response to aselected pathogen. For example, a serology test may be conducted todetect antibodies to selected pathogens in a patient's sera. Inconnection with this aspect of the invention, antigenic determinants ofa selected pathogen may be chosen for use in an immunogenic compositionon a selected patient based on a diagnostic indication that the patienthas had one or more prior exposure(s) to the pathogen, for example byvirtue of the presence of antibodies to antigenic determinants of thatpathogen in the patient's sera.

In further selected embodiments, the invention involves diagnostic stepsto assess a patient's immunological response to treatment with aselected immunogenic composition. For example, the diagnostic steps mayinclude evaluating a patient's immune response to the antigenicdeterminants of that immunogenic composition, for example using aserological test to detect antibodies to those antigenic determinants.In connection with this aspect of the invention a treatment with aselected immunogenic composition may be continued if the evaluationindicates that there is an active immunological response to theantigenic determinants of that composition, and the treatment may bediscontinued, and an alternative treatment with a different immunogeniccomposition may be initiated, if the evaluation indicates that there isnot a sufficiently active immunological response to the antigenicdeterminants of the immunogenic composition.

One aspect of the invention involves the treatment of pathologiesassociated with microbial lung infections with antigenic compositionsthat comprise antigenic determinants of microbial pathogens that areknown to be lung pathogens, such as exogenous lung pathogens orpathogens that are members of the endogenous flora of the respiratorysystem. For example, antigenic determinants of the endogenous bacterialrespiratory flora species that most commonly cause infection in the lung(see Table 5) may be used to treat infections situated in the lung:Streptococcus pneumoniae, Moraxella catarrhalis, Mycoplasma pneumoniae,Klebsiella pneumoniae, Haemophilus influenza. Similarly, common virallung pathogens from Table 5 may be selected for use in some embodiments.Alternatively, a more exhaustive list of endogenous lung pathogens maybe selected from Table 1, based on the pathogenicity informationprovided in Table 2. In further alternative embodiments, viral lungpathogens listed in Table 4 may be used. And in further alternativeembodiments, exogenous bacterial lung pathogens from Table 3 may be usedin formulating antigenic compositions of the invention, i.e. selectedfrom the group consisting of: Achromobacter spp., Actinomadura spp.,Alcaligenes spp., Anaplasma spp., Bacillus anthracis, other Bacillusspp., Balneatrix spp., Bartonella henselae, Bergeyella zoohelcum,Bordetella holmesii, Bordetella parapertussis, Bordetella pertussis,Borrelia burgdorferi, Borrelia recurrentis, Brucella spp., Burkholderiagladioli, Burkholderia mallei, Burkholderia pseudomallei, Campylobacterfetus, Capnoctyophaga canimorsus, Capnoctyophaga cynodegmi, Chlamydiapneumoniae, Chlamydia psittaci, Chlamydophila pneumoniae,Chromobacterium violaceum, Chlamydophila psittaci, Chryseobacteriumspp., Corynebacterium pseudotuberculosis, Coxiella burnetii, Francisellatularensis, Gordonia spp., Legionella spp., Leptospirosis spp.,Mycobacterium avium, Mycobacterium kansasii, Mycobacterium tuberculosis,other Mycobacterium spp., Nocardia spp., Orientia tsutsugamushi,Pandoraea spp., Pseudomonas aeruginosa, other Pseudomonas spp.,Rhodococcus spp., Rickettsia conorii, Rickettsia prowazekii, Rickettsiarickettsiae, Rickettsia typhi.

Infections may also arise in bronchial tissue and therefore, in someembodiments, antigenic compositions that comprise antigenic determinantsof microbial pathogens that are known to cause bronchial infection maybe used to treat patients with infections situated in the bronchialtissue, including, for example, the following common causes of bronchialinfection: Mycoplasma pneumoniae, Chlamydophila pneumoniae, Bordetellapertussis, Streptococcus pneumoniae, Haemophilus influenzae, influenzavirus, adenovirus, rhinovirus, coronavirus, parainfluenza, respiratorysyncytial virus, human metapneumovirus, or coxsackievirus. Infectionsthat are located in both lung and bronchial tissue may be treated withantigenic compositions that comprise antigenic determinants of microbialpathogens that are known to cause both lung and bronchial infection (forexample, Streptococcus pneumoniae, Haemophilus influenza and Mycoplasmapneumoniae are all common lung and bronchial pathogens) oralternatively, with antigenic compositions that comprise antigenicdeterminants of microbial pathogens that are known to cause lunginfection and antigenic determinants of microbial pathogens that areknown to cause bronchial infection.

One aspect of the invention involves the treatment of pathologiesassociated with microbial infections of the colon with antigeniccompositions that comprise antigenic determinants of heterologousmicrobial pathogens that are known to be colon pathogens, such aspathogens that are members of the endogenous flora of the colon orexogenous colonic pathogens. For example, antigenic determinants of thefollowing microbial species may be used to treat heterologous infectionssituated in the colon: Escherichia coli, Clostridium difficile,Bacteroides fragilis, Bacteroides vulgatus, Bacteroidesthetaiotaomicron, Clostridium perfringens, Salmonella enteriditis,Yersinia enterocolitica, Shigella flexneri; adenoviruses, astroviruses,caliciviruses, noroviruses, rotaviruses, or cytomegalovirus. In selectedembodiments, antigenic determinants of E. coli, the most commonbacterial cause of colon infection, may be used alone or with antigenicdeterminants of other common pathogens of the colon to treat pathologiesassociated with infections of the colon, such as pathologies associatedwith infections caused by heterologous strains of E. coli.

In alternative aspects, the invention utilizes microbial antigens, suchas bacterial or viral antigens, to formulate antigenic compositions,where the microbial species is selected on the basis of the tissue ororgan within which the microbe is known to cause infections. Bacterialresident flora are the most common bacterial pathogens, accounting forthe vast majority of bacterial infections in most animals, includinghumans. Resident flora can for example infect through primaryattachment, or attachment and invasion following mucosa damage,resulting for example from vascular, trauma, chemical insult, or damageresulting from primary infection.

For microbial pathogens, virulence and infection potential is acombination of the ability of the microbe to adhere, to produce enzymes,to survive immunoproducts (complement, antibody) and to survive themicrobiocidal activity of macrophage and neutrophils. Some bacteria,including endogenous bacteria, may be sufficiently virulent as to causemonomicrobial infections, while others are more effective with thesynergy of polymicrobial infection. In general, it is often not possibleto be precise about the specific role of individual microbes within themilieu of mixed infection. As acute infection may, in some cases,provide more optimal immune stimulation, accordingly, in someembodiments, the invention utilizes microbial species that are involvedin acute infection.

In some embodiments, bacteria that are members of the endogenous floraof a particular region may be used to formulate antigenic compositionsof the invention. The rows of Table 6 list a number of bacterialspecies, together with the biological regions in which each species mayform a part of the endogenous flora. For example, Abiotrophia spp. aretypically members of the endogenous flora of the respiratory tract andthe mouth. Further and for example, the organisms listed in Table 6 maybe used as microbial pathogens to formulate antigenic compositions, orantigenic compositions having those pathogens may be selected, for useto treat heterologous infections, for example as an anti-microbialtreatment for heterologous infections situated in the tissues or organslisted with the relevant organism in Table 6.

TABLE 6 Human Bacterial Normal Flora (Endogenous Bacterial HumanPathogens) Duodenum/ GU Bacterial species Respiratory Mouth StomachJejunum Ileum Colon System Genital Skin CFU /mL 10{circumflex over ( )}510{circumflex over ( )}2 10{circumflex over ( )}5 10{circumflex over( )}8 10{circumflex over ( )}11 Abiotrophia + + spp Acholeplasma + +laidlawii Acidaminococcus + + + + + fermentansAcinetobacter + + + + + + + + spp. Actinobacillus + + spp.Actinobaculum + + + + + spp. Actinomyces + + + + + + + spp. Aerococcus +christensenii Aerococcus + viridans Aerococcus + urinae Aeromonas + + +spp. Alloiococcus + otitis Anaerorhabdus + + furcosusAnaerococcus + + + + hydrogenalis Anaerococcus + + + lactolyticusAnaerococcus + + + prevotii Arcanobacterium + + spp. Atopobium + + + + +spp. Bacillus + + + spp. Bacteroides + + caccae Bacteroides + +distasonis Bacteroides + + eggerthii Bacteroides + + + + fragilisBacteroides + + merdae Bacteroides + + ovatus Bacteroides + +splanchnicus Bacteroides + + thetaiotaomicron Bacteroides + + vulgatusBifidobacterium + + + adolescentis Bifidobacterium + + + + + bifidumBifidobacterium + + + + + breve Bifidobacterium + + + + + catenulatumBifidobacterium + + + + + + + dentium Bifidobacterium + + + + + longumBilophila + + + + + + + wadsworthia Brevibacterium + caseiBrevibacterium + epidermidis Burkholderia + + + + cepaciaButyrivibrio + + + fibrisolvens Campylobacter + + + + concisusCampylobacter + + + + curvus Campylobacter + + + + gracilisCampylobacter + + + jejuni Campylobacter + + + rectusCampylobacter + + + + + showae Campylobacter + + sputorumCapnocytophaga + + granulosum Capnocytophaga + + gingivalisCampylobacter + + haemolytica Capnocytophaga + + + + + + + ochraceaCapnocytophaga + + sputigena Cardiobacterium + + hominis Cedecea + sppCentipeda + + periodontii Citrobacter + + + freundii Citrobacter + + +koseri Clostridium + + + spp. Corynebacterium + + + accolensCorynebacterium + + + afermentans Corynebacterium + amycolatumCorynebacterium + auris Corynebacterium + + diphtheriaeCorynebacterium + durum Corynebacterium + glucuronolyticumCorynebacterium + jeikeium Corynebacterium + macginleyiCorynebacterium + matruchotii Corynebacterium + minutissimumCorynebacterium + propinquum Corynebacterium + pseudodiphtheriticumCorynebacterium + riegelii Corynebacterium + simulansCorynebacterium + + striatum Corynebacterium + ulceransCorynebacterium + + urealyticum Dermabacter + hominis Dermacoccus +nishinomiyaensis Desulfomonas + + + pigra Dysgonomonas + + + spp.Eikenella + + + + + corrodens Enterobacter + + + aerogenesEnterobacter + + + cloacae Enterobacter + + + gergoviaeEnterobacter + + + sakazakii Enterobacter + + + tayloraeEnterococcus + + + spp. Escherichia + + + + + coli Escherichia + + +fergusonii Escherichia + + + hermannii Escherichia + + + vulnerisEubacterium + + + + + spp. Ewingella + + americana Finegoldia + + + + +magnus Fusobacterium + + alocis Fusobacterium + + + + + gonidiaformansFusobacterium + + + mortiferum Fusobacterium + + + + + naviformeFusobacterium + + + + + necrophorum Fusobacterium + + + nucleatumFusobacterium + + sulci Fusobacterium + + + russii Fusobacterium + + +varium Gardnerella + + + + + vaginalis Gemella + + haemolysansGemella + + + + + morbillorum Globicatella + + spp. Granulicatella + +spp. Haemophilus + + + spp. Hafnia alvei + + + Helcococcus + kunziiHelicobacter + + + spp. Kingella + + spp. Klebsiella + + + + + spp.Kocuria + spp. Kytococcus + sedentarius Lactobacillus + + + + + + + +acidophilus Lactobacillus + + breve Lactobacillus + + + + caseiLactobacillus + + cellobiosus Lactobacillus + + + + + + + + fermentumLactobacillus + + + + reuteri Lactobacillus + + + + + + salivariusLactococcus + + spp. Leclercia + + + adecarboxylata Leminorella + + +spp. Leptotrichia + + + + buccalis Leuconostoc + + spp.Megasphaera + + + elsdenii Micrococcus + + + luteus Micrococcus + + +lylae Micromonas + + micros Mitsuokella + + + multiacidusMobiluncus + + + + curisii Mobiluncus + + + + mulieris Moellerella + + +wisconsensis Moraxella + + catarrhalis other + + + + Moraxella spp.Morganella + + + morganii Mycoplasma + + buccale Mycoplasma + fauciumMycoplasma + + + fermentans Mycoplasma + + genitalium Mycoplasma + + +hominis Mycoplasma + + lipophilum Mycoplasma + + orale Mycoplasma +penetrans Mycoplasma + + pneumoniae Mycoplasma + primatum Mycoplasma + +salivarium Mycoplasma + spermatophilum Neisseria + cinerea Neisseria +flavescens Neisseria + lactamica Neisseria + + meningitidis Neisseria +mucosa Neisseria + polysaccharea Neisseria + sicca Neisseria + subflavaOligella + + ureolytica Oligella + + urethralis Pantoea + + +agglomerans Pastuerella + + bettyae Pasteurella + + multocidaPediococcus + + spp. Peptococcus + + + niger Peptoniphilus + + + + + +asaccharolyticus Peptoniphilus + lacrimalis Peptostreptococcus + + + + +anaerobus Peptostreptococcus + + + productus Peptostreptococcus + + +vaginalis Porphyromonas + + + + + + asaccharolytica Porphyromonas + + +catoniae Porphyromonas + + + endodontalis Porphyromonas + + + gingivalisPrevotella + + bivia Prevotella + + + buccae Prevotella + + + + +buccalis Prevotella + + + corporis Prevotella + + + dentalisPrevotella + + + denticola Prevotella + + disiens Prevotella + + +enoeca Prevotella + + + heparinolytica Prevotella + + + intermediaPrevotella + + + + + loescheii Prevotella + + + + + melaninogenicaPrevotella + + + nigrescens Prevotella + + + + + oralis Prevotella + + +oris Prevotella + + + oulorum Prevotella + + + tanneraePrevotella + + + + + veroralis Prevotella + + + zoogleoformansPropionibacterium + acnes Propionibacterium + avidum Propionibacterium +granulosum Propionibacterium + + propionicum Propionferax + innocuumProteus + + + mirabilis Proteus + + + penneri Proteus + + + vulgarisProvidencia + + rettgeri Providencia + + + stuartii Pseudomonas + + +aeruginosa Retortamonas + + + intestinalis Rothia + + dentocariosaRothia + + mucilaginosa Ruminococcus + + + productus Selenomonas + +spp. Serratia + + liquefaciens Serratia + + marcescens Serratia + +odorifera Staphylococcus + + + + + aureus Staphylococcus + auricularisStaphylococcus + capitis Staphylococcus + caprae Staphylococcus + cohniiStaphylococcus + + + + + epidermidis Staphylococcus + haemolyticusStaphylococcus + hominis Staphylococcus + lugdunensis Staphylococcus +pasteuri Staphylococcus + saccharolyticus Staphylococcus + +saprophyticus Staphylococcus + schleiferia Staphylococcus + simulansStaphylococcus + xylosus Staphylococcus + warneriStreptococcus + + + + + agalactiae Streptococcus + + + + + + + anginosusStreptococcus + + + bovis Streptococcus + + + + + + + constellatusStreptococcus + + criceti Streptococcus + + crista Streptococcus + +equisimilis Streptococcus + + gordonii Streptococcus + + + + + +intermedius Streptococcus + + + mitis Streptococcus + + mutansStreptococcus + + oralis Streptococcus + + parasanguis Streptococcus +pneumoniae Streptococcus + + + + pyogenes Streptococcus + + + salivariusStreptococcus + + + sanguis Streptococcus + + sobrinus Streptococcus + +vestibularis Group C + G + + Streptococci Succinivibrio + + +dextrinosolvens Sutterella + + + + + spp. Suttonella + + indologenesTissierella + + + praeacuta Treponema + + denticola Treponema + +maltophilum Treponema + minutum Treponema + phagedenis Treponema +refringens Treponema + + socranskii Treponema + + vincentii Turicella +otitidis Ureaplasma + + + urealyticum Veillonella + + + + + spp.Weeksella + + virosa

Endogenous microbial flora, such as bacteria, have access to tissues forpathogenesis either through contiguous spread or bacteremic spread.Under favorable conditions, all endogenous organisms can becomepathogenic and invade locally and spread by contiguous spread toadjacent tissues and organs. Endogenous bacterial flora of the skin,mouth and colon are the species that are understood to also be amenableto bacteremic spread. Bacteria that are members of a particularendogenous flora domain may therefore cause infection in tissues ororgans to which these bacteria may spread. Accordingly, one aspect ofthe invention involves the use of endogenous microbial pathogens totreat an infection of a tissue or organ to which the endogenous bacteriamay spread to cause infection. The columns of Table 7 list 9 domains forendogenous flora, the: skin, respiratory system, genitals, GU system,mouth, stomach, duodenum/jejunum, ileum and colon. The rows of Table 7list organs or tissues within which heterologous microbial infectionsmay be situated. Accordingly, one aspect of the invention involves theuse of endogenous microbial pathogens to formulate antigeniccompositions, or the selection of existing formulations having thepathogens, for treating heterologous microbial infections situated intissues or organs to which the pathogen may spread to cause aninfection. Accordingly, in alternative embodiments, infections situatedin the tissues or organs listed in the first column of Table 7 may betreated with antigenic compositions comprising antigenic determinantsthat are specific for microbial pathogens that are members of theendogenous flora of one or more of the endogenous flora domains listedin the first row of Table 7 and indicated with an X or a check mark inthe appropriate row. For example, infections situated in the prostatemay be treated with an antigenic composition having antigenicdeterminants specific for a microbial pathogen or pathogens endogenousto the GU system and/or genital system. A number of the bacterialspecies that are endogenous to the endogenous flora domains listed inTable 7 are listed, with the corresponding endogenous flora domains, inTable 6. Accordingly, one aspect of the invention involves the treatmentof an infection situated in a tissue listed in Table 7 with an antigeniccomposition comprising antigenic determinants of the bacterial speciesthat are listed in Table 6, where the regions of endogenous flora linkedto the site of the infection in Table 7 match the regions of endogenousflora linked to the bacterial species in Table 6. The examples providedin Tables 6 and 7 may be used to formulate antigenic compositions foruse in treating heterologous microbial infections, for example as ananti-microbial treatment for heterologous microbial infections in theorgans identified in Tables 6 and 7.

TABLE 7 Tissue/Organ Pathogenicity of Endogenous Flora Tissue/organ GUDuodenum/ site Skin Respiratory Genital System Mouth Stomach JejunumIleum Colon Skin X X Soft tissue X (i.e. fat and muscle) (e.g., sarcoma)Breast X X Lymph nodes: X X X head and neck Lymph nodes: X ✓ ✓axillae/arm Lymph nodes: X ✓ ✓ mediastinal Lymph nodes: X pulmonaryhilum Lymph nodes: X ✓ X X X X intra- abdominal Lymph nodes: X X ✓ ✓inguinal/leg Hematological ✓ ✓ ✓ (e.g. leukemias, multiple myeloma) BoneX ✓ ✓ Joints X ✓ ✓ ✓ Meninges X X Brain ✓ ✓ ✓ Spinal cord ✓ ✓ ✓Eye/Orbit X X X X Salivary X glands Oral X Tonsil X X Nasopharynx/ X XSinus Thyroid ✓ ✓ ✓ Larynx X X Lung/Trachea/ X Bronchi Pleura ✓ X ✓ ✓ ✓Mediastinum X Heart ✓ ✓ □ Esophagus X Stomach X Small bowel X XColon/Rectum X Anus X X Perineum X X Liver ✓ ✓ ✓ Gallbladder X Biliarytract X Pancreas X Spleen ✓ ✓ ✓ Adrenal gland ✓ ✓ ✓ Kidney ✓ X ✓ ✓Ureter X Bladder ✓ X X Peritoneum X X X X Retroperitoneal X X X X X areaProstate X X Testicle X X Penis X X X Ovary/Adnexae X X X Uterus X X XCervix X X X Vagina X X Vulva X X * Bacteria have access totissues/organs either through: Contiguous spread (X) or Bacteremicspread: (✓).

In accordance with the combined information in Tables 6 and 7,infections located in the tissues or organs set out in column 1 of Table7 may be treated with antigenic compositions comprising antigenicdeterminants of the corresponding but heterologous bacterial species ofTable 6, so that the column headings in Table 7 are in effect replacedwith the bacterial species of Table 6.

In some embodiments, microbial pathogens for use in the invention may beexongenous bacterial pathogens. For example, the organisms listed inTable 8 may be used as microbial pathogens to formulate antigeniccompositions, or antigenic compositions having those pathogens may beselected, for use to treat pathologies associated with heterologousinfections situated in the tissues or organs listed with the relevantorganism in Table 8. In some embodiments, antigenic determinants of bothendogenous and exogenous bacterial species targeted to a specific tissueor organ may be used in combination. For example, an antigeniccomposition derived from, or specific for, Clostridium difficile, may beused to treat pathologies associated with heterologous microbialinfections in the colon.

TABLE 8 Exogenous Bacterial Human Pathogens, and their Sites ofInfection bacterial species tissue/organ sites Achromobacterhematological, skin, soft tissue, spp. lung/trachea/bronchi, peritoneum,meninges, bile duct, gallbladder, kidney, bladder, ureter Actinomaduraskin, soft tissue, lung/trachea/ spp. bronchi, mediastinum, brain,spinal cord, hematological, meninges, joints Aerobacter spp. smallbowel, colon, hematological, Aerococcus spp. peritoneum hematological,heart, bone, kidney, bladder, ureter, meninges Alcaligenes spp.lung/trachea/bronchi Anaplasma spp. meninges, hematological, liver,spleen, bone, lung/trachea/bronchi Bacillus anthracislung/trachea/bronchi, lymph nodes pulmonary hilum, mediastinum,meninges, skin, nasopharynx, tonsil, oral, small bowel, colon,hematological Bacillus cereus colon, eye, hematological other Bacillushematological, bone, meninges, brain, heart, spp. lung/trachea/bronchi,mediastinum, skin, soft tissue, colon, stomach, small bowel, eyeBalneatrix spp. lung/trachea/bronchi, meninges, hematological Bartonellaskin, hematological, liver, muscle, bacilliformis lymph nodes, jointsBartonella brain, spinal cord, hematological, henselae skin, liver,bone, pleura, lung/trachea/bronchi, mediastinum, axillary and inguinallymph nodes, eye, joints Bartonella skin, hematological, liver, spleen,joints quintana Bergeyella skin, soft tissue, meninges, hematological,zoohelcum lung/trachea/bronchi Bordetella lung/trachea/bronchi,hematological holmesii Bordetella nasopharynx, tonsil,lung/trachea/bronchi parapertussis Bordetella nasopharynx, tonsil,lung/trachea/bronchi pertussis Borrelia meninges, brain, spinal cord,burgdorferi skin, eye, hematological, inguinal/axillary/cervical lymphnodes, muscle, liver, spleen, nasopharynx, lung/trachea/ bronchi,testes, joints Borrelia brain, spinal cord, hematological, recurrentissmall bowel, liver, spleen, salivary glands, lung/trachea/bronchi, lymphnodes, eye, skin Brevundimonas peritoneum, hematological, skin, softtissue spp. Brucella spp. lung/trachea/bronchi, lymph nodes pulmonaryhilum, meninges, brain, spinal cord, lymph nodes, mediastinum, bone,eye, small bowel, colon, liver, biliary tract, kidney, ureter, bladder,hematological, skin, testes, spleen, prostate, joints Burkholderiahematological, meninges, gladioli lung/trachea/bronchi Burkholderialung/trachea/bronchi, skin, mallei soft tissue, liver, spleen, muscle,lymph nodes pulmonary hilum, mediastinal lymph nodes, mediastinum, headand neck lymph nodes, hematological Burkholderia lung/trachea/bronchi,skin, pseudomallei kidney, bladder, ureter, soft tissue, bone, brain,spinal cord, muscle, hematological, prostate, kidney, ureter, meningesCalymmatobacterium skin, penis, vulva, soft tissue, vagina, cervix,granulomatis bone, hematological, inguinal lymph nodes Campylobactersmall bowel, colon coli Campylobacter lung/trachea/bronchi, small bowel,fetus colon, meninges, brain, peritoneum, bone, gallbladder, ovaries,hematological, heart, kidney, bladder, ureter Campylobacter colon,hematological, gallbladder, pancreas, jejuni bladder, bone, meningesCampylobacter small bowel, colon sputorum Capnoctyophaga skin, softtissue, meninges, hematological, bone, canimorsus lung/trachea/bronchi,eye Capnoctyophaga skin, soft tissue, meninges, hematological, bone,cynodegmi lung/trachea/bronchi, eye CDC groups EF- hematological, eye,skin, soft tissue 4a and EF-4b Chlamydia lung/trachea/bronchi, lymphnodes pneumoniae pulmonary hilum, liver, brain, meninges, skin, thyroid,pancreas, hematological Chlamydia psittaci lung/trachea/bronchi, lymphnodes pulmonary hilum, mediastinum, liver, brain, meninges,hematological, skin, thyroid, pancreas Chlamydia inguinal lymph nodes,penis, trachomatis vulva, vagina, cervix, uterus, ovaries and adnexae,peritoneum, prostate, eye Chlamydophila laryngx, trachea/bronchi,hematological pneumoniae Chromobacterium hematological, liver, spleen,violaceum lung/trachea/bronchi, kidney, bladder, ureter, eye/orbit,bone, brain, meninges, spinal cord Chlamydophila lung/trachea/bronchipsittaci Chryseobacterium meninges, lung/trachea/bronchi, hematologicalspp. Clostridium small bowel, colon, stomach, skin, soft tissue,bifermentans hematological Clostridium colon, small bowel, skinbotulinum Clostridium colon difficile Clostridium indolis small bowel,colon, stomach, skin, soft tissue, hematological Clostridium smallbowel, colon, stomach, skin, soft tissue, mangenolii hematologicalClostridium small bowel, colon, stomach, skin, soft tissue, perfringenshematological, heart Clostridium small bowel, colon, stomach, skin, softtissue, sordellii hematological Clostridium small bowel, colon, stomach,skin, soft tissue, sporogenes hematological Clostridium small bowel,colon, stomach, skin, soft tissue, subterminale hematologicalClostridium tetani skin, soft tissue Comamonas spp. hematological,peritoneum, eye Corynebacterium neck/axillary/inguinal/mediastinalpseudotuberculosis lymph nodes, lymph nodes pulmonary hilum,lung/trachea/bronchi, mediastinum Coxiella bumetii lung/bronchi/trachea,brain, spinal cord, liver, bone, joints Edwarsiella tarda skin, softtissue, liver, meninges, small bowel, colon, bone, uterus, ovariesEhrlichia spp. meninges, brain, spinal cord, hematological, bone, liver,kidney, spleen, lymph nodes Erysipelothrix skin, hematological, bone,brain, peritoneum rhusiopathiae Francisella nasopharynx, oral, tonsil,tularensis lung/trachea/bronchi, skin, axillary/head and neck/inguinallymph nodes, hematological, eye, small bowel Fusobacterium skin, softtissue, hematological spp. Gordonia spp. skin, soft tissue,lung/trachea/ bronchi, mediastinum, brain, spinal cord, hematological,meninges, eye Haemophilus skin, inguinal lymph nodes, penis, vulva,vagina ducreyi Helicobacter stomach pylori Legionella spp.lung/trachea/bronchi, lymph nodes pulmonary hilum, hematological, brain,spinal cord, muscle, pancreas Leptospirosis spp. lung/trachea/bronchi,pancreas, meninges, brain, spinal cord, skin, lymph nodes, eye,hematological, nasopharynx, oral, tonsil, kidney, liver, spleen ListeriaHematological (septicemia), monocytogenes brain, meninges, spinal cord,small bowel, colon, GIT, cornea, lung, uterous or cervix in pregnantwomen Methylobacterium hematological, peritoneum, skin, spp. softtissue, bone Mycobacterium lung/bronchi/trachea, lymph avium nodespulmonary hilum, prostate, pancreas, spleen, skin, neck lymph nodes,esophagus, bone, hematological Mycobacterium colon, small bowel, jointsbovis Mycobacterium lung/bronchi/trachea, lymph nodes kansasii pulmonaryhilum, prostate, bone Mycobacterium skin, soft tissues, testes, eyeleprae Mycobacterium skin, soft tissue, bone marinum Mycobacterium headand neck lymph nodes scrofulaceum Mycobacterium lung/bronchi/trachea,lymph tuberculosis nodes pulmonary hilum, prostate, peritoneum,pancreas, spleen, lymph nodes, small bowel, meninges, brain, spinalcord, kidney, ureter, bladder, muscle, esophagus, colon, testes, eye,ovaries, cervix, vagina, uterus, mediastinum, larynx, skin,hematological, pleura, joints Mycobacterium skin, soft tissue ulceransother lung/bronchi/trachea, lymph nodes Mycobacterium pulmonary hilum,skin, spp. soft tissues, bone, head and neck lymph nodes, jointsMyroides spp. kidney, bladder, ureter, skin, soft tissue, hematologicalNeisseria nasopharyx, oral, tonsil, prostate, gonorrhoeae penis, vagina,cervix, uterus, ovary/adnexae, peritoneum, skin, muscle, bone, liver,hematological, head and neck and inguinal and intra- abdominal lymphnodes, anus, joints Neorickettsia hematological, bone, lymph nodes,liver, spleen sennetsu Nocardia spp. lung/bronchi/trachea, pancreas,meninges, spinal cord, brain, skin, soft tissue, eye, bone, kidney,heart, hematological Orientia meninges, brain, spinal cord,tsutsugamushi hematological, skin, inguinal and axillary lymph nodes,spleen, lung/bronchi/trachea Pandoraea spp. lung/trachea/bronchi,hematological Pasteurella canis skin, soft tissue, hematologicalPasteurella skin, soft tissue, hematological dagmatis Pasteurella skin,soft tissue, hematological stomatis Pediococcus spp. hematological,liver, colon Pityrosporum skin ovale Plesiomonas small bowel, colon,hematological, shigelloides meninges, bone, gall bladder, skin, softtissue Pseudomonas lung/trachea/bronchi, hemaotogical, aeruginosa skin,soft tissue, bone, meninges, brain, eye, kidney, bladder, ureter, heart,joints other skin, soft tissue, lung/trachea/bronchi, Pseudomonasmediastinum, hematological spp. Ralstonia spp. hematological, meninges,bone Rhizobium spp. hematological, peritoneum, eye, kidney, bladder,ureter Rhodococcus lung/trachea/bronchi, hematological, spp. brain,skin, lymph nodes, bone, mediastinum, liver, spleen, soft tissue, spinalcord, meninges Rickettsia akari skin Rickettsia conoriilung/bronchi/trachea, lymph nodes pulmonary hilum, meninges, brain,spinal cord, hematolofical, skin, kidney, liver, spleen, pancreasRickettsia felis skin, brain, spinal cord Rickettsia meninges, brain,spinal cord, hematological, prowazekii lung/bronchi/trachea, skin,spleen Rickettsia lung/bronchi/trachea, lymph rickettsiae nodespulmonary hilum, meninges, brain, spinal cord, hematological, muscle,small bowel, liver, skin Rickettsia slovaca skin, head and neck lymphnodes Rickettsia typhi meninges, hematological, liver, kidney, brain,lung/bronchi/trachea, spleen Roseomonas spp. hematological, peritoneum,skin, soft tissue, bladder, kidney, ureter Salmonella spp.lung/bronchi/trachea, pancreas, spleen, intra-abdominal lymph nodes,stomach, small bowel, colon, meninges, skin, muscle, bone,hematological, heart, joints Shewanella spp. skin, soft tissue, eye,bone, hematological, peritoneum Shigella boydii colon Shigella colondysenteriae Shigella flexneri colon Shigella sonnei colonSphingobacterium brain, meninges, spinal cord, eye, spp. skin, softtissue Sphingomonas hematological, meninges, peritoneum, skin, spp. softtissue, kidney, bladder, ureter Spirillum minus skin,axillary/inguinal/neck lymph nodes, hematological, liver, spleen otherSpirillum colon spp. Stenotrophomonas meninges, hematological,peritoneum, maltophilia lung/trachea/bronchi, eye, kidney, bladder,ureter, skin, soft tissue Streptobacillus skin, bone, hematological,moniliformis lung/trachea/bronchi, meninges, brain, liver, spleenStreptococcus skin, hematological, soft tissue iniae Streptococcus smallbowel, nasopharynx, bone, meninges, zooepidemicus hematological, headand neck lymph nodes Streptomices spp. skin, soft tissue, lung/trachea/bronchi, mediastinum, brain, spinal cord, hematological, meninges,joints Treponema nasopharynx, tonsil, oral, pallidum meninges, brain,spinal cord, penis, vulva, vagina, anus, cervix, eye, hematological,inguinal and head and neck lymph nodes Tropheryma brain, spinal cord,hematological, whipplei small bowel, colon, heart, lung/trachea/bronchi,eye Tsukamurella skin, soft tissue, lung/trachea/ spp. bronchi,mediastinum, brain, spinal cord, hematological, meninges Vibrio choleraecolon, small bowel Vibrio hematological, meninges cincinnatiensis Vibriodamsela skin, soft tissue Vibrio fluvialis small bowel, colon Vibriofumissii small bowel, colon Vibrio hollisae small bowel, colon, skin,soft tissue Vibrio hematological metschnikovii Vibrio colon, small bowelparahaemolyticus Vibrio vulnificus soft tissue, blood, skin Yersinianasopharynx, tonsil, small bowel, enterocolitica intra-abdominal lymphnodes, colon, muscle, lung/trachea/bronchi, liver, spleen,hematological, joints Yersinia pestis lung/trachea/bronchi, lymph nodespulmonary hilum, inguinal/axillary/neck lymph nodes, oral, tonsil,hematological, skin, joints Yersinia small bowel, colon, abdominalpseudotuberculosis lymph nodes, joints

In some embodiments, microbial pathogens for use in the invention may beviral pathogens. Table 9 provides an exemplary list of viral pathogenstogether with the tissue and organ sites for which each viral species isunderstood to be a pathogen. Accordingly, one aspect of the inventioninvolves utilizing immunogenic compositions that are specific for thenamed viruses to treat pathologies associated with infections caused byheterologous micro-organisms in the organs or tissues that areidentified adjacent to the name of the virus in Table 9. For example, anantigenic composition derived from, or specific for, a vaccinia virus,may be used to treat a condition characterized by an infection by aheterologous micro-organism in the skin, hematological tissues, lymphnodes, brain, spinal cord, eye or heart.

TABLE 9 Viral Human Pathogens and Their Sites of Infection virustissue/organ sites Vaccinia skin, hematological, lymph nodes, brain,spinal cord, eye, heart Variola (smallpox) skin, hematological, lymphnodes, brain Monkeypox skin, hematological, head and neck lymph nodes,brain, eye, lung/trachea/bronchi, pulmonary hilar lymph nodes,mediastinum, nasopharynx Cowpox skin, hematological, lymph nodesParapoxviruses Skin Molluscum skin contagiosum Tanapox skin,hematological, axillary and inguinal lymph nodes Herpes Simplexnasopharynx, oral, tonsil, hematological, virus (1 and 2)lung/bronchi/trachea, pancreas, meninges, brain, spinal cord, inguinaland head/neck lymph nodes, penis, vulva, perineum, esophagus, liver,eye, skin, rectum, tonsil, mediastinum, anus, vagina, cervixVaricella-zoster nasopharynx, sinus, lung/ trachea/bronchi, pulmonaryhilar lymph nodes, hematological, pancreas, meninges, brain, spinalcord, esophagus, liver, eye, skin, heart, mediastinum Cytomegalovirusnasopharynx, lymph nodes, tonsil, hematological, lung/trachea/bronchi,pancreas, abdominal lymph nodes, brain, spinal cord, esophagus, smallbowel, colon/rectum, eye, liver, heart, skin, mediastinum, esophagusEpstein-Barr virus nasopharynx, tonsil, oral, lymph nodes,hematological, lung, abdominal lymph nodes, brain, spinal cord, muscles,esophagus, liver, heart, skin, spleen, kidney, muscle, heart,lung/trachea/ bronchi, pulmonary hilar lymph nodes, mediastinum Humanherpesvirus skin, hematological, lung/ 6 trachea/bronchi, pulmonaryhilar lymph nodes, brain, meninges, liver Human herpesvirus skin, brain,liver 7 Human herpesvirus nasopharynx, tonsil, hematological, 8 skin,spleen, head and neck lymph nodes Simian herpes B brain, spinal cord,skin, hematological, virus lymph nodes Adenovirus nasopharynx, oral,larynx, trachea, bronchi, lung, lymph nodes, meninges, brain, spinalcord, small bowel, colon, liver, intra-abdominal lymph nodes,mediastinum, bladder, sinus, hematological, ureter, kidney, bladder,thyroid, heart BK virus kidney Human skin, anus, penis, vulva, cervix,papillomavirus vagina, oral Hepatitis B virus liver, pancreas,hematological, bone, joints Hepatitis D virus liver Parvovirus B19 skin,hematological, nasopharynx, bone, kidney, heart, liver, brain, meninges,joints Orthoreoviruses nasopharynx, small bowel, colon, oral, sinus,lymph nodes, skin, lung/trachea/ bronchi, meninges, brain, spinal cord,liver Orbiviruses brain, muscle, hematological, Coltiviruseshematological, skin, muscle, oral, spleen, lymph nodes, meninges, brainRotaviruses small bowel, colon, liver, hematological, pancreas,nasopharynx, billiary tract, meninges, brain Alphaviruses brain, spinalcord, small bowel, colon, hematological, skin, bone Rubella skin,hematological, head and neck lymph nodes, spleen, nasopharynx, bone,brain, tonsil, bronchi, liver, heart, joints Yellow fever virushematological, liver, lung/ trachea/bronchi, kidney, adrenal gland,spleen, lymph nodes, stomach, kidney Dengue fever virus hematological,lymph nodes, skin, spleen, muscle, liver, brain, nasopharynx, jointsJapanese brain, hematological, spinal cord encephalitis virus West Nilebrain, hematological, spinal encephalitis virus cord, muscle, lymphnodes, liver, spleen, pancreas, meninges St. Louis brain, hematological,spinal cord, encephalitis virus meninges, muscle, nasopharynx Tick-bornebrain, hematological, spinal cord, encephalitis virus muscle, meningesother Flaviviruses hematological, brain, meninges, bone, muscles, skin,lymph nodes Hepatitis C virus hematological, liver, joints Hepatitis Gvirus liver Coronaviruses nasopharynx, sinus, oral, tonsil, larynx,lung/trachea/bronchi, pulmonary hilar lymph nodes, small bowel, colon,tonsil, hematological Toroviruses small bowel, colon, hematologicalParainfluenza nasopharynx, sinus, tonsil, oral, larynx, viruseslung/trachea/bronchi, pulmonary hilar lymph nodes, meninges,hematological, mediastinum Mumps virus salivary glands, pancreas, brain,spinal cord, liver, testes, hematological, meninges, ovaries, bone,heart, kidney, thyroid, prostate, breast, joints Respiratorynasopharynx, tonsil, sinus, syncytial lung/trachea/bronchi, viruspulmonary hilar lymph nodes, mediastinum, hematological, oral, pleuraHuman nasopharynx, lung/trachea/ metapneumovirus bronchi, pulmonaryhilar lymph nodes, tonsil, sinus, mediastinum, hematological, oral,pleura, larynx, eye, skin, small bowel, colon Rubeola nasopharynx,sinus, hematological, lung/trachea/bronchi, pulmonary hilar lymph nodes,intra-abdominal lymph nodes, meninges, brain, spinal cord, liver,spleen, lymph nodes, skin, thymus, eye, oral, heart Hendra virus brain,meninges, lung/trachea/bronchi, kidney, hematological, muscle, Nipahvirus brain, meninges, spleen, lymph nodes, thymus,lung/trachea/bronchi, kidneys, brain, spinal cord, meninges,hematological Vesicular hematological, muscle, oral, stomatitis tonsil,nasopharyngeal, virus lymph nodes, small bowel, colon Rabies virus skin,meninges, brain, spinal cord, oral, nasopharynx, salivary glands,hematological Lyssaviruses brain, spinal cord Influenza virusnasopharynx, laryngx, lung/ trachea/bronchi, pulmonary hilar lymphnodes, meninges, muscle, hematological, mediastinum, muscle, sinus,tonsil, oral, eye, pleura, brain, spinal cord, salivary glands, thyroid,heart California hematological, brain, meninges encephalitis virusHantaviruses hematological, kidney, eye, skin, oral, muscle,lung/trachea/bronchi other Bunyaviruses brain, hematological, muscle,meninges, spinal cord Lymphocytic hematological, muscle, lymph nodes,skin, brain, choriomeningitis meninges, testes, bone virus Lassa virusnasopharynx, brain, spinal cord, lung/trachea/bronchi, pulmonary hilarlymph nodes, mediastinum, muscle, testes, eye, heart, Machupo virusbrain, meninges, hematological, muscle, eye, skin, lymph nodes,nasopharynx, small bowel, colon Junin virus brain, meninges,hematological, muscle, eye, skin, lymph nodes, nasopharynx, small bowel,colon Human T-Cell hematological, skin, lymph Lymphotropic nodes,muscle, eye, bone, viruses lung, pulmonary hilar lymph nodes, spinalcord, brain Poliovirus nasopharynx, lung/trachea/ bronchi, pulmonaryhilar lymph nodes, small bowel, neck and intra-abdominal lymph nodes,colon, hematological, liver, spleen, skin, brain, spinal cord, meninges,heart Coxsackieviruses nasopharynx, larynx, oral, tonsil,lung/trachea/bronchi, pulmonary hilar lymph nodes, mediastinum,pancreas, muscle, brain, meninges, small bowel, neck and intra-abdominal lymph nodes, colon, hematological, spleen, skin, eye, sinus,liver, testes, bone, pleura, salivary glands, heart Echovirusesnasopharynx, oral, tonsil, lung/trachea/bronchi, pulmonary hilar lymphnodes, muscle, brain, meninges, small bowel, neck and intra-abdominallymph nodes, colon, hematological, mediastinum, spleen, skin, eye,sinus, liver, pancreas, testes, bone, salivary glands, heart otherlung/trachea/bronchi, pulmonary hilar lymph Enteroviruses nodes,meninges, brain, skin, heart, joints Hepatitis A virus small bowel,colon, hematological, liver, spleen, brain, spinal cord, gallbladder,pancreas, kidney Rhinoviruses nasopharynx, sinus, oral, tonsil, larynx,lung/trachea/bronchi, pulmonary hilar lymph nodes Noroviruses and smallbowel, colon other Caliciviruses Astroviruses small bowel, colonPicobirnaviruses small bowel, colon Hepatitis E virus liver, smallbowel, colon, hematological

The cumulative information in Tables 6 through 9 provides an extensiveidentification of microbial pathogens that may be used in theformulation of antigenic compositions of the invention, together with anidentification of the tissues or organs in which these organisms arepathogenic, and accordingly identifies the correspondence betweenselected tissues or organs in which an infection by a heterologousorganism is situated, and the organisms that may be used to produceantigenic formulations for treating the condition.

In some embodiments, the microbial pathogen selected for use inantigenic compositions of the invention may be one that is a commoncause of acute infection in the tissue or organ in which theheterologous infection is to be treated. Table 10 identifies bacterialand viral pathogens of this kind, together with the tissues and organsin which they commonly cause infection. Accordingly, in selectedembodiments, an infection residing in a tissue identified in the firstcolumn of Table 10 may be treated with an antigenic composition thatcomprises antigenic determinants for one or more of the heterologouspathogenic organisms listed in the second column of Table 10. Forexample, an infection in the skin may be treated with an antigeniccomposition comprising antigenic determinants of one or more of thefollowing heterologous organisms: Staphylococcus aureus, Beta hemolyticstreptococci group A, B, C and G, Corynebacterium diptheriae,Corynebacterium ulcerans, Pseudomonas aeruginosa, rubeola, rubella,varicella-zoster, echoviruses, coxsackieviruses, adenovirus, vaccinia,herpes simplex, or parvo B19.

TABLE 10 Common Causes of Acute Infection (Bacterial and Viruses) ForEach Tissue/Organ Site Tissue/organ Common Bacterial or Viral Pathogensof specific site tissue/organ site Skin Staphylococcus aureus, Betahemolytic streptococci group A, B, C and G, Corynebacterium diptheriae,Corynebacterium ulcerans, Pseudomonas aeruginosa rubeola, rubella,varicella-zoster, echoviruses, coxsackieviruses, adenovirus, vaccinia,herpes simplex, parvo B19 Soft tissue (i.e. Streptococcus pyogenes,Staphylococcus aureus, fat and muscle) Clostridium perfringens, otherClostridium spp. (e.g., sarcoma) influenza, coxsackieviruses BreastStaphylococcus aureus, Streptococcus pyogenes Lymph nodes:Staphylococcus aureus, Streptococcus pyogenes head and neckEpstein-Barr, cytomegalovirus, adenovirus, measles, rubella, herpessimplex, coxsackieviruses, varicella-zoster Lymph nodes: Staphylococcusaureus, Streptococcus pyogenes axillae/arm measles, rubella,Epstein-Barr, cytomegalovirus, adenovirus, varicella-zoster Lymph nodes:viridans streptococci, Peptococcus spp., mediastinal Peptostreptococcusspp., Bacteroides spp., Fusobacterium spp., Mycobacterium tuberculosismeasles, rubella, Epstein-Barr, cytomegalovirus, varicella- zoster,adenovirus Lymph nodes: Streptococcus pneumoniae, Moraxella catarrhalis,pulmonary Mycoplasma pneumoniae, Klebsiella pneumoniae, hilumHaemophilus influenza, Chlamydophila pneumoniae, Bordetella pertussis,Mycobacterium tuberculosis influenza, adenovirus, rhinovirus,coronavirus, parainfluenza, respiratory syncytial virus, humanmetapneumovirus, coxsackievirus Lymph nodes: Yersinia enterocolitica,Yersinia pseudotuberculosis, intra-abdominal Salmonella spp.,Streptococcus pyogenes, Escherichia coli, Staphylococcus aureus,Mycobacterium tuberculosis measles, rubella, Epstein-Barr,cytomegalovirus, varicella- zoster, adenovirus, influenza,coxsackieviruses Lymph nodes: Staphylococcus aureus, Streptococcuspyogenes inguinal/leg measles, rubella, Epstein-Barr, cytomegalovirus,herpes simplex Hematological Staphylococcus aureus, Streptococcuspyogenes, (e.g. leukemias, coagulase-negative staphylococci,Enterococcus spp., multiple myeloma) Escherichia coli, Klebsiella spp.,Enterobacter spp., Proteus spp., Pseudomonas aeruginosa, Bacteroidesfragilis, Streptococcus pneumoniae, group B streptococci rubeola,rubella, varicella-zoster, echoviruses, coxsackieviruses, adenovirus,Epstein-Barr, cytomegalovirus, herpes simplex Bone Staphylococcusaureus, coagulase-negative staphylococci, Streptococcus pyogenes,Streptococcus pneumoniae, Streptococcus agalactiae, other streptococcispp., Escherichia coli, Pseudomonas spp., Enterobacter spp., Proteusspp., Serratia spp. parvo virus B19, rubella, hepatitis B JointStaphylococcus aureus, coagulase-negative staphylococci, Streptococcuspyogenes, Streptococcus pneumoniae, Streptococcus agalactiae, otherstreptococci spp., Escherichia coli, Pseudomonas spp., Enterobacterspp., Proteus spp., Serratia spp., Neisseria gonorrhea, salmonellaspecies, Mycobacterim tuberculosis, Hemophilus influenza parvo virusB19, rubella, hepatitis B Scedosporium prolificans Meninges Haemophilusinfluenzae, Neisseria meningitidis, Streptococcus pneumoniae,Streptococcus agalactiae, Listeria monocytogenes echoviruses,coxsackieviruses, other enteroviruses, mumps Brain Streptococcus spp.(including S. anginosus, S. constellatus, S. intermedius),Staphylococcus aureus, Bacteroides spp., Prevotella spp., Proteus spp.,Escherichia coli, Klebsiella spp., Pseudomonas spp., Enterobacter spp.,Borrelia burgdorferi coxsackieviruses, echoviruses, poliovirus, otherenteroviruses, mumps, herpes simplex, varicella-zoster, flaviviruses,bunyaviruses Spinal cord Haemophilus influenzae, Neisseria meningitidis,Streptococcus pneumoniae, Streptococcus agalactiae, Listeriamonocytogenes, Borrelia burgdorferi coxsackieviruses, echoviruses,poliovirus, other enteroviruses, mumps, herpes simplex,varicella-zoster, flaviviruses, bunyaviruses Eye/Orbit Staphylococcusaureus, Streptococcus pyogenes, Streptococcus pneumoniae, Streptococcusmilleri, Escherichia coli, Bacillus cereus, Chlamydia trachomatis,Haemophilus influenza, Pseudomonas spp., Klebsiella spp., Treponemapallidum adenoviruses, herpes simplex, varicella-zoster, cytomegalovirusSalivary glands Staphylococcus aureus, viridans streptococci (e.g.,Streptococcus salivarius, Streptococcus sanguis, Streptococcus mutans),Peptostreptococcus spp., Bacteroides spp., and other oral anaerobesmumps, influenza, enteroviruses, rabies Oral Prevotella melaninogenicus,anaerobic streptococci, viridans streptococci, Actinomyces spp.,Peptostreptococcus spp., Bacteroides spp., and other oral anaerobesherpes simplex, coxsackieviruses, Epstein-Barr Tonsil Streptococcuspyogenes, Group C and G B-hemolytic streptococci rhinoviruses,influenza, coronavirus, adenovirus, parainfluenza, respiratory syncytialvirus, herpes simplex Sinus Streptococcus pneumoniae, Haemophilusinfluenza, Moraxella catarrhalis, α-streptococci, anaerobic bacteria(e.g., Prevotella spp.), Staphylococcus aureus rhinoviruses, influenza,adenovirus, parainfluenza Nasopharynx Streptococcus pyogenes, Group Cand G B-hemolytic streptococci rhinoviruses, influenza, coronavirus,adenovirus, parainfluenza, respiratory syncytial virus, herpes simplexThyroid Staphylococcus aureus, Streptococcus pyogenes, Streptococcuspneumoniae mumps, influenza Larynx Mycoplasma pneumoniae, Chlamydophilapneumoniae, Streptococcus pyogenes rhinovirus, influenza, parainfluenza,adenovirus, corona virus, human metapneumovirus Trachea Mycoplasmapneumoniae parainfluenza, influenza, respiratory syncytial virus,adenovirus Bronchi Mycoplasma pneumoniae, Chlamydophila pneumoniae,Bordetella pertussis, Streptococcus pneumoniae, Haemophilus influenzaeinfluenza, adenovirus, rhinovirus, coronavirus, parainfluenza,respiratory syncytial virus, human metapneumovirus, coxsackievirus LungStreptococcus pneumoniae, Moraxella catarrhalis, Mycoplasma pneumoniae,Klebsiella pneumoniae, Haemophilus influenza influenza, adenovirus,respiratory syncytial virus, parainfluenza Pleura Staphylococcus aureus,Streptococcus pyogenes, Streptococcus pneumoniae, Haemophilusinfluenzae, Bacteroides fragilis, Prevotella spp., Fusobacteriumnucleatum, peptostreptococcus spp., Mycobacterium tuberculosisinfluenza, adenovirus, respiratory syncytial virus, parainfluenzaMediastinum viridans streptococci, Peptococcus spp., Peptostreptococcusspp., Bacteroides spp., Fusobacterium spp. measles, rubella,Epstein-Barr, cytomegalovirus Heart Streptococcus spp. (including S.mitior, S. bovis, S. sanguis, S. mutans, S. anginosus), Enterococcusspp., Staphylococcus spp., Corynebacterium diptheriae, Clostridiumperfringens, Neisseria meningitidis, Salmonella spp. enteroviruses,coxsackieviruses, echoviruses, poliovirus, adenovirus, mumps, rubeola,influenza Esophagus Actinomyces spp., Mycobacterium avium, Mycobacteriumtuberculosis, Streptococcus spp. cytomegalovirus, herpes simplex,varicella-zoster Stomach Streptococcus pyogenes, Helicobacter pyloricytomegalovirus, herpes simplex, Epstein-Barr, rotaviruses, noroviruses,adenoviruses Small bowel Escherichia coli, Clostridium difficile,Bacteroides fragilis, Bacteroides vulgatus, Bacteroidesthetaiotaomicron, Clostridium perfringens, Salmonella enteriditis,Yersinia enterocolitica, Shigella flexneri adenoviruses, astroviruses,caliciviruses, noroviruses, rotaviruses, cytomegalovirus Colon/RectumEscherichia coli, Clostridium difficile, Bacteroides fragilis,Bacteroides vulgatus, Bacteroides thetaiotaomicron, Clostridiumperfringens, Salmonella enteriditis, Yersinia enterocolitica, Shigellaflexneri adenoviruses, astroviruses, caliciviruses, noroviruses,rotaviruses, cytomegalovirus Anus Streptococcus pyogenes, Bacteroidesspp., Fusobacterium spp., anaerobic streptococci, Clostridium spp., E.coli, Enterobacter spp., Pseudomonas aeruginosa, Treponema pallidumherpes simplex Perineum Escherichia coli, Klebsiella spp., Enterococcusspp., Bacteroides spp., Fusobacterium spp., Clostridium spp.,Pseudomonas aeruginosa, anaerobic streptococci, Clostridium spp., E.coli, Enterobacter spp. herpes simplex Liver Escherichia coli,Klebsiella spp., Streptococcus (anginosus group), Enterococcus spp.,other viridans streptococci, Bacteroides spp. hepatitis A, Epstein-Barr,herpes simplex, mumps, rubella, rubeola, varicella-zoster,coxsackieviruses, adenovirus Gallbladder Escherichia coli, Klebsiellaspp., Enterobacter spp., enterococci, Bacteroides spp., Fusobacteriumspp., Clostridium spp., Salmonella enteriditis, Yersinia enterocolitica,Shigella flexneri Biliary tract Escherichia coli, Klebsiella spp.,Enterobacter spp., Enterococci spp., Bacteroides spp., Fusobacteriumspp., Clostridium spp., Salmonella enteriditis, Yersinia enterocolitica,Shigella flexneri hepatitis A, Epstein-Barr, herpes simplex, mumps,rubella, rubeola, varicella-zoster, cocsackieviruses, adenovirusPancreas Escherichia coli, Klebsiella spp., Enterococcus spp.,Pseudomonas spp., Staphylococcal spp., Mycoplasma spp., Salmonellatyphi, Leptospirosis spp., Legionella spp. mumps, coxsackievirus,hepatitis B, cytomegalovirus, herpes simplex 2, varicella-zoster SpleenStreptococcus spp., Staphylococcus spp., Salmonella spp., Pseudomonasspp., Escherichia coli, Enterococcus spp. Epstein-Barr, cytomegalovirus,adenovirus, measles, rubella, coxsackieviruses, varicella-zoster Adrenalgland Streptococcus spp., Staphylococcus spp., Salmonella spp.,Pseudomonas spp., Escherichia coli, Enterococcus spp. varicella-zosterKidney Escherichia coli, Proteus mirabilis, Proteus vulgatus,Providentia spp., Morganella spp., Enterococcus faecalis, Pseudomonasaeruginosa BK virus, mumps Ureter Escherichia coli, Proteus mirabilis,Proteus vulgatus, Providentia spp., Morganella spp., Enterococcus spp.Bladder Escherichia coli, Proteus mirabilis, Proteus vulgatus,Providentia spp., Morganella spp., Enterococcus faecalis,Corynebacterium jekeum adenovirus, cytomegalovirus PeritoneumStaphylococcus aureus, Streptococcus pyogenes, Streptococcus pneumonia,Escherichia coli, Klebsiella spp., Proteus spp., Enterococci spp.,Bacteroides fragilis, Prevotella melaninogenica, Peptococcus spp.,Peptostreptococcus spp., Fusobacterium spp., Clostridium spp.Retroperitoneal Escherichia coli, Staphylococcus aureus area ProstateEscherichia coli, Klebsiella spp., Enterobacter spp., Proteus mirabilis,Enterococci spp., Pseudomonas spp., Corynebacterium spp., Neisseriagonorrhoeae herpes simplex Testicle Escherichia coli, Klebsiellapneumoniae, Pseudomonas aeruginosa, Staphylococcus spp., Streptococcusspp., Salmonella enteriditis mumps, coxsackievirus, lymphocyticchoriomeningitis virus Penis Staphylococcus aureus, Streptococcuspyogenes, Neisseria gonorrhoeae, Treponema pallidum herpes simplex,human papillomavirus Ovary/Adnexae Neisseria gonorrhoeae, Chlamydiatrachomatis, Gardenerella vaginalis, Prevotella spp., Bacteroides spp.,Peptococcus spp. Streptococcus spp., Escherichia coli Uterus Neisseriagonorrhoeae, Chlamydia trachomatis, Gardenerella vaginalis, Prevotellaspp., Bacteroides spp., Peptococcus spp., Streptococcus spp.,Escherichia coli Cervix Neisseria gonorrhoeae, Chlamydia trachomatis,Treponema pallidum herpes simplex Vagina Gardenerella vaginalis,Prevotella spp., Bacteroides spp., peptococci spp., Escherichia coli,Neisseria gonorrhoeae, Chlamydia Trachomatis, Treponema pallidum, herpessimplex Vulva Staphylococcus aureus, Streptococcus pyogenes, Treponemapallidum herpes simplex

In selected embodiments, particular microbial pathogens are suited fortreatment of particular pathologies associated with heterologousmicrobal infections located in the tissue or organ within which theorganism is pathogenic, examples of selected embodiments are set out inTable 10. These are exemplary embodiments, and not an exhaustive list ofthe alternative formulations for use in accordance with the invention.

The specific microbes which commonly cause infection in a specifictissue or organ may vary by geographical location. For example,Mycobacterium tuberculosis is a more common cause of lung infection insome geographical locations and populations than in others andtherefore, while M. tuberculosis may not be a common lung pathogen insome geographic and population groups it may be a common lung pathogenin others. Table 10 is thus not an exhaustive list of common pathogensfor all geographic locations and population groups. It is understoodthat a clinical microbiologist skilled in the art could determine thecommon pathogenic species in a particular geographic area or populationgroup for a specific tissue or organ site in accordance with theinvention. For veterinary use, there will of course be specificpathogens that are common in selected tissues of selected species, andthis may also vary geographically.

In selected embodiments, the invention involves diagnostic steps toassess a patient's previous exposure to microbial pathogens. Forexample, the diagnostic steps may include taking a medical history ofexposure to selected pathogens, and/or evaluating a patient's immuneresponse to a selected pathogen. For example, a serology test may beconducted to detect antibodies to selected pathogens in a patient'ssera. In connection with this aspect of the invention, antigenicdeterminants of a selected microbial pathogen may be chosen for use inan immunogenic composition on a selected patient based on a diagnosticindication that the patient has had one or more prior exposure(s) to thepathogen, for example by virtue of the presence of antibodies toantigenic determinants of that pathogen in the patient's sera.

In further selected embodiments, the invention involves diagnostic stepsto assess a patient's immunological response to treatment with aselected immunogenic composition. For example, the diagnostic steps mayinclude evaluating a patient's immune response to the antigenicdeterminants of that immunogenic composition, for example using aserological test to detect antibodies to those antigenic determinants.In connection with this aspect of the invention a treatment with aselected immunogenic composition may be continued if the evaluationindicates that there is an active immunological response to theantigenic determinants of that composition, and the treatment may bediscontinued, and an alternative treatment with a different immunogeniccomposition may be initiated, if the evaluation indicates that there isnot a sufficiently active immunological response to the antigenicdeterminants of the immunogenic composition.

In selected embodiments, the microbial pathogen selected for use inantigenic compositions of the invention may be one that is the mostcommon cause of acute infection in the tissue or organ in which theheterologous infection is to be treated. For example, for the treatmentof pathologies associated with infections of the bone, Staphylococcusaureus would be the bacterial species selected for treatment ofinfections caused by heterologous organisms; for the treatment ofinfections in lung tissue, Streptococcus pneumoniae would be selectedfor treatment of infections caused by heterologous organisms; for thetreatment of breast infections, Staphylococcus aureus would be selectedfor treatment of infections caused by heterologous organisms; for thetreatment of kidney or bladder infections, Escherichia coli would beselected for treatment of infections caused by heterologous organisms;and for the treatment of infections in the colon, Escherichia coli wouldbe the bacterial species selected for treatment of infections caused byheterologous organisms. It is understood that a clinical microbiologistskilled in the art could determine the most frequently pathogenicspecies, bacterial or viral, for each specific tissue or organ inaccordance with the invention. In selected embodiments, only antigenicdeterminants of the most common pathogen for the particular tissue ororgan are used to treat heterologous infections of that tissue or organ.In alternative embodiments, antigenic determinants of the most commonpathogen for the particular tissue or organ could be used in combinationwith antigenic determinants of other pathogens that are known to bepathogenic in the of that particular tissue or organ, preferentiallyselecting from the more common pathogens.

In some embodiments, the invention provides antigenic compositions inwhich a threshold proportion of antigenic determinants selected inaccordance with the invention are used, relative to any other antigenicdeterminants in the composition. For example, antigenic compositions mayhave greater than X % of the antigenic determinants therein derived frompathogenic (or commonly pathogenic, or most commonly pathogenic)species, where X may for example be 10, 30, 40, 50, 60, 70, 80, 90, 95or 100 (or any integer value between 10 and 100). For example, at leastX % of the antigenic determinants in the antigenic composition may bespecific for microbial pathogens that are pathogenic (or commonlypathogenic or most commonly pathogenic) in the specific organ or tissueof the patient within which the heterologous infection is situated.Using an alternative measure, of the total number of microbial pathogensin the antigenic composition, at least X % may be selected to bemicrobial pathogens that are pathogenic (or commonly pathogenic or mostcommonly pathogenic) in the specific organ or tissue of the patientwithin which the heterologous microbial infection is situated. In someembodiments, the antigenic composition may accordingly consistessentially of antigenic determinants of one or more microbial pathogensthat are each pathogenic (or commonly pathogenic or most commonlypathogenic) in the specific organ or tissue of the patient within whichthe heterologous infection is situated.

In some embodiments, the invention comprises the use of bacterial orviral vaccines or formulations that are approved for other purposes(e.g., poliomyelitis vaccine, H. influenza vaccine, meningococcalvaccine, pneumococcal vaccine, influenza vaccine, hepatitis B vaccine,hepatitis A vaccine, diphtheria vaccine, tetanus vaccine, pertussisvaccine, measles vaccine, mumps vaccine, rubella vaccine, varicellavaccine, BCG vaccine, cholera vaccine, Japanese encephalitis vaccine,rabies vaccine, typhoid vaccine, yellow fever vaccine, small poxvaccine, etc.) for use as treatments of infections caused byheterologous micro-organisms by selecting a vaccine containing apathogen (or antigenic constituent of a pathogen) that is pathogenic inthe specific organ or tissue of the patient within which theheterologous infection is situated by consulting Tables 6-10. Forexample, a S. pneumoniae vaccine, either a whole cell vaccine or avaccine comprised of one or more antigenic components of S. pneumoniae(e.g., pneumococcal polysaccharide-23-valent) could be used to treat aheterologous infection at any of the following sites in which S.pneumoniae is listed as a common pathogen in Table 10: pulmonary hilarlymph nodes, bone, meninges, spinal cord, eye/orbit, sinus, thyroid,bronchi, lungs, pleura or peritoneum. As a further example, a hepatitisB vaccine could be used to treat a heterologous infection at any of thefollowing sites in which hepatitis B virus is listed as a pathogen inTable 9, as follows: liver, pancreas, or hematological infections.

In some embodiments, selected compositions and methods are specificallyexcluded from the scope of the invention. For example, the use of the aformulation of antigens of a particular microbial pathogen in thetreatment of a pathology associated with infection by that organism. Forexample, selected embodiments exclude the use of PVF or MRV vaccines forthe treatment of lung infections caused by the organisms that arepresent in those formulations.

Example 1: Murine Studies

Example 1a: Illustrating the influence of a heat inactivated Klebsiellapneumoniae antigenic composition on monocyte/macrophage and dendriticcell populations in mice.

The following methods and materials were utilized in this Example:

Mice. C57BL/6 female mice 7-8 weeks of age were ordered from Harlan Labs(Livermore, Calif.) for these studies.

Antibodies and reagents. The following antibodies were used in thisExample: anti-I-A/I-E FITC (MHC Class II; M5/114.15.2); anti-Gr-1 PE(RB6-8C5); anti-CD11 b PerCP-Cy5 (M1/70); anti-CD11c APC (N418);anti-CD4 FITC (GK1.5); anti-NK1.1 PE (PK136); anti-CD8a eFluor780(53-6.7); anti-CD44 APC (IM7). All antibodies were acquired fromeBioscience (San Diego, Calif.). Liberase™ and DNAse I was acquired fromRoche. All media was from HyClone (Fisher).

Treatment with antigenic compositions. Heat killed K. pneumoniae withphenol (KO12 [5.0 OD600 units]) was diluted 1/10 in PBS containing 0.4%phenol and 100 μl was injected subcutaneously on day 0, 2, 4, and 6 into4 mice. Control mice (n=5) were injected on day 0, 2, 4, and 6 with PBS.

Brochoalveolar lavage. On day 7 mice were sacrificed and abronchoalveolar lavage (BAL) was performed by exposing the tracheafollowed by insertion of a 22G catheter attached to a 1 ml syringe. 1 mlof PBS was injected into the lungs and removed and placed into a 1.5 mlmicrocentrifuge tube. The lungs were subsequently washed 3 more timeswith 1 ml of PBS and the fluid was pooled. The first wash from eachmouse was centrifuged at 400×g and the supernatant was frozen forcytokine analysis. The final 3 ml of lavage fluid was centrifuged andthe cells were pooled with the cell pellet from the first lavage. Thecells were counted and stained with antibodies specific for MHC class11, Ly6G/C, CD11b, and CD11c. After staining the cells were washed andanalyzed on a FACS Calibur flow cytometer.

Lung digestion. After BAL was performed the lungs were placed in 5 ml ofRPMI containing 417.5 μg/ml Liberase TL (Roche) and 200 μg/ml DNAse I(Roche). The lungs were then digested at 37° C. for 30 mins. Afterdigestion the lungs were forced through a 70 um cell strainer to createa single cell suspension. The cells were then centrifuged, washed,resuspended in FACS Buffer (PBS with 2% FCS and 5 mM EDTA) and counted.After counting the cells were stained and analyzed by FACS using thesame antibodies as for the BAL cells.

Peritoneal lavage. 1 ml of PBS was injected into the peritoneum of miceusing a 1 ml syringe attached to a 25G needle after BAL. The abdomen wasmassaged for 1 minute and 0.5 ml of PBS was recovered from theperitoneum using a 1 ml pipet. The lavage fluid was put in a 1.5 mlcentrifuge tube, centrifuged at 400×g for 5 mins, and resuspended inFACS buffer prior to staining and FACS analysis.

Spleen and lymph node analysis. The spleen and draining lymph node wereremoved after BAL and peritoneal lavage and placed in PBS. The spleenwas disrupted by mashing through a 70 μm cell strainer (Fisher) and thelymph node was disrupted using the rubber end of the plunger from a 1 mlsyringe. After disruption, the single cell suspension from the spleenand lymph nodes was centrifuged, washed once with FACS Buffer, andresuspended in FACS Buffer prior to counting, staining, and FACSanalysis.

FAGS Analysis. Cells were stained on ice for 20 mins in 96 well platesusing 50 ul of antibodies diluted in FACS buffer. After 20 mins, 100 μlof FACs buffer was added to the wells and the plates were centrifuged at400×g for 5 mins. Subsequently the media was removed and the cells werewashed 1 more time with FACS buffer. After the final wash the cells wereresuspended in 200 μl of FACS buffer and the data was acquired using aFACS Calibur flow cytometer (BD). A minimum of 20,000 live events werecollected for all samples except the BAL where a minimum of 5,000 eventswas collected.

The following results were obtained in this Example.

Normal mice were treated with a K. pneumoniae antigenic composition onday 0, 2, 4, and 6. On day 7 the mice were sacrificed and thebronchoalveolar lavage fluid, lung tissue, peritoneal lavage fluid,lymph nodes, and spleen was analyzed for changes in monocyte andmacrophages. An increase in the number of acute inflammatory bloodmonocytes/macrophages, defined by high expression of CD11b and Gr-1(same marker as Ly6c), and F4/80 in the lymph node draining the site ofinjection of the K. pneumoniae antigenic composition was observed (see:FIG. 1A). These acute inflammatory monocytes/macrophages also expressvery high levels of MHC class II molecules suggesting exposure tobacterial antigens. Importantly, treatment of mice with the K.pneumoniae antigenic composition for one week led to a marked increasein the frequency of acute inflammatory monocytes in the bronchoalveolarlavage fluid and in the lungs (i.e., the targeted organ) but not in thespleen or peritoneum of treated mice, suggesting that treatment caninduce homing of monocytes specifically to the lungs without affectingother organs (see: FIG. 1B). Monocytes can differentiate into dendriticcells (DCs) in the lungs and consistent with our observations of amarked increase in monocyte recruitment it was also observed that therewas a marked increase in the frequency of cells displaying markers formature DCs (see: FIG. 1C).

As illustrated in FIGS. 1A to 1C, treatment with a K. pneumoniaeantigenic composition for 7 days resulted in a marked increase (comparedto treatment with placebo=PBS) in both acute inflammatory monocytes anddendritic cells in the lungs of mice. As illustrated in FIGS. 1A to 1C,mice were treated with either a K. pneumoniae antigenic composition foror PBS on day 0, 2, 4, and 6. On day 7, the mice were sacrificed and thetotal number of A) and B) inflammatory monocytes (CD11b+ Gr-1+ cells)and C) dendritic cells (CD11c+ MHC class II+ cells) were determined byflow cytometry in the lung and spleen. The error bars depicted in A)represent the mean of 4-5 mice per group.

Example 1B. Illustrating the influence of a heat inactivated Klebsiellapneumoniae antigenic composition and a heat inactivated E. coliantigenic composition on monocyte/macrophage, dendritic cell, andeffector cell populations in mice

The following methods and materials were utilized in this Example:

Mice. C57BL/6 female mice 7-8 weeks of age were ordered from Harlan Labs(Livermore, Calif.) for these studies.

Antibodies and reagents. The following antibodies were used:anti-1-A/I-E FITC (MHC Class II; M5/114.15.2); anti-Gr-1 PE (RB6-8C5);anti-CD11 b PerCP-Cy5 (M1/70); anti-CD11c APC (N418); anti-CD4 FITC(GK1.5); anti-NK1.1 PE (PK136); anti-CD8a eFluor780 (53-6.7); anti-CD44APC (IM7). All antibodies were acquired from eBioscience (San Diego,Calif.). Liberase™ and DNAse I was acquired from Roche. All media wasfrom HyClone (Fisher).

Treatment with antigenic compositions. Heat-killed K. pneumoniae withphenol (K. pneumoniae; lot KO12; 5.0 OD600 units) was diluted 1/10 inPBS containing 0.4% phenol and 100 ul was injected subcutaneously on day0, 2, 4, and 6 into 5 mice. Heat-killed E. coli (lot; 5.0 OD600 units)was diluted 1/10 in containing 0.4% phenol and 100 μl was injectedsubcutaneously on day 0, 2, 4, and 6 into 5 mice. Control mice (n=5)were injected on day 0, 2, 4, and 6 with PBS.

Brochoalveolar lavage. On day 7 mice were sacrificed and abronchoalveolar lavage (BAL) was performed by exposing the tracheafollowed by insertion of a 22G catheter attached to a 1 ml syringe. 1 mlof PBS was injected into the lungs and removed and placed into a 1.5 mlmicrocentrifuge tube. The lungs were subsequently washed 3 more timeswith 1 ml of PBS and the fluid was pooled. The first wash from eachmouse was centrifuged at 400×g and the supernatant was frozen forcytokine analysis. The final 3 ml of lavage fluid was centrifuged andthe cells were pooled with the cell pellet from the first lavage. Thecells were counted and stained with antibodies specific for MHC classII, Ly6G/C, CD11 b, and CD11c. After staining the cells were washed andanalyzed on a FACS Calibur flow cytometer.

Lung digestion. After BAL was performed the lungs were placed in 5 ml ofRPMI containing 417.5 μg/ml Liberase TL (Roche) and 200 μg/ml DNAse I(Roche). The lungs were then digested at 37° C. for 30 mins. Afterdigestion the lungs were forced through a 70 μm cell strainer to createa single cell suspension. The cells were then centrifuged, washed,resuspended in FACS Buffer (PBS with 2% FCS and 5 mM EDTA) and counted.After counting the cells were stained and analyzed by FACS using thesame antibodies as for the BAL cells.

Peritoneal lavage. 1 ml of PBS was injected into the peritoneum of miceusing a 1 ml syringe attached to a 25G needle after BAL. The abdomen wasmassaged for 1 minute and 0.5 ml of PBS was recovered from theperitoneum using a 1 ml pipet. The lavage fluid was put in a 1.5 mlcentrifuge tube, centrifuged at 400×g for 5 mins, and resuspended inFACS buffer prior to staining and FACS analysis.

Spleen and lymph node analysis. The spleen and draining lymph node wereremoved after BAL and peritoneal lavage and placed in PBS. The spleenwas disrupted by mashing through a 70 μm cell strainer (Fisher) and thelymph node was disrupted using the rubber end of the plunger from a 1 mlsyringe. After disruption, the single cell suspension from the spleenand lymph nodes was centrifuged, washed once with FACS Buffer, andresuspended in FACS Buffer prior to counting, staining, and FACSanalysis.

FAGS Analysis. Cells were stained on ice for 20 mins in 96 well platesusing 50 μl of antibodies diluted in FACS buffer. After 20 mins, 100 μlof FACs buffer was added to the wells and the plates were centrifuged at400×g for 5 mins. Subsequently the media was removed and the cells werewashed 1 more time with FACS buffer. After the final wash the cells wereresuspended in 200 μl of FACS buffer and the data was acquired using aFACS Calibur flow cytometer (BD). A minimum of 20,000 live events werecollected for all samples except the BAL where a minimum of 5,000 eventswas collected.

The following results were obtained in this Example:

As illustrated in FIG. 2, mice were treated on day 0, 2, 4, and 6 witheither a K. pneumoniae antigenic composition, an E. coli antigeniccomposition or PBS. On day 7 the mice were sacrificed and the totalnumber of inflammatory monocytes (CD11b+ Gr-1+ cells) and dendriticcells (CD11c+ MHC class II+ cells) were determined by flow cytometry inthe peritoneal lavage fluid, lungs, lymph node and spleen. Error bars inFIG. 18 represent the standard deviation from 5 mice. *p-value<0.05using a Student's t-test.

FIG. 2 illustrates that treatment with a K. pneumoniae antigeniccomposition, but not an E. coli antigenic composition treatment,markedly increased the number of monocytes and DCs in the lungs of mice.In contrast to the lungs, K. pneumoniae did not lead to an increase inmonocytes in the peritoneum of the mice whereas E. coli did.Importantly, there was only a slight increase in the number ofinflammatory monocytes and no increase in DCs in the spleens of micetreated with either K. pneumoniae or E. coli suggesting that the effectsof the therapies are not general and are, in fact, specific for aparticular organ site. In addition to looking at the effects oftreatment on inflammatory monocytes and DCs in the lungs of mice, wealso looked at changes in other leukocytes such as cytotoxic CD8 Tcells, CD4 T helper cells, and natural killer (NK) cells.

FIG. 3 illustrates that a K. pneumoniae antigenic composition, but notPBS or an E. coli antigenic composition, resulted in a marked increasein the frequency and total numbers of NK cells, CD4 and CD8 T cells inthe lungs of treated mice. This Example demonstrates that subcutaneousinjection of a killed bacterial species which normally causes lunginfection can promote the accumulation of leukocytes in the lungswithout the presence of any inflammation in that site. In addition, itdemonstrates that this effect is specific to the targeted site and thatit is also specific to the bacterial constituents of the treatment used.

As illustrated in FIG. 3, mice were treated on day 0, 2, 4, and 6 witheither a K. pneumoniae antigenic composition, an E. coli antigeniccomposition, or PBS. On day 7, the mice were sacrificed and the totalnumber of CD4 T cells, CD8 T cells, and natural killer (NK) cells weredetermined by flow cytometry. Error bars represent the sd of valuesobtained from 5 mice per group. *p-value<0.05 using a Student's t-test.

Example 1C. Illustrating the effects of heat, irradiation, and phenolinactivation on K. pneumoniae antigenic compositions, includingleukocyte recruitment into the lungs of mice, and the effects of phenolas a preservative.

The following methods and materials were utilized in this Example:

Mice. C57BL/6 female mice 7-8 weeks of age were ordered from Harlan Labs(Livermore, Calif.) for these studies.

Antigenic compositions. Heat killed K. pneumoniae antigenic compositionwith phenol (KO12), heat killed K. pneumoniae antigenic compositionwithout phenol (KO25), irradiated K. pneumoniae antigenic compositionwithout phenol (KO24), and phenol killed K. pneumoniae antigeniccomposition without phenol (KO25) were used in this study. All bacterialformulations were at a concentration of 5.0 OD units in saline. For 1/10dilution, 1 ml of bacterial formulation was added to 9 ml of DPBS andmixed immediately and then again prior to injection. For 1/100 dilution,0.1 ml of bacterial formulation will be added to 9.9 ml of DPBS andmixed immediately and then again prior to injection. For dilutions ofheat-killed Klebsiella pneumoniae antigenic composition with phenol, thedilutions were carried out as above using a DPBS solution containing0.4% phenol (w/v). To prepare the 0.4% phenol in DPBS, first a 5% phenolsolution was prepared by adding 0.5 g of solid phenol (Sigma Aldrich,St. Louis, Mo.) to 10 ml of DPBS (Hyclone, Logan, Utah) This solutionwas filtered through a 0.22 um filter (Millipore, Billerica, Mass.) andstored at 4° C. Immediately prior to use the 5% phenol solution wasdiluted 1 ml in 12.5 ml DPBS and used to prepare the bacterialformulations.

Treatment with antigenic compositions. 5 mice per group were treatedsubcutaneously on day 0, 2, 4, and 6 with 0.1 ml of a heat-killed K.pneumoniae antigenic composition diluted 1/10 in PBS or PBS with 0.4%phenol, 0.1 ml of an irradiated K. pneumoniae antigenic compositiondiluted 1/10 in PBS, or a phenol inactivated K. pneumoniae antigeniccomposition diluted 1/10 with PBS or PBS with 0.4% phenol. On day 7 themice were sacrificed and leukocyte recruitment to the lungs was analyzedas in Example 1B.

The following results were obtained in this Example:

In this example, we used leukocyte recruitment to the lungs as asurrogate of efficacy to compare the efficacy of K. pneumoniae antigeniccompositions inactivated by various methods. FIGS. 4A to 4B illustratethat, for both heat killed and phenol killed K. pneumoniae antigeniccompositions, the addition of phenol (0.4%) as a preservative, increasedefficacy, as measured by cellular recruitment. In some embodiments, asmall amount of phenol (i.e., 0.4% as a preservative) may stabilize acomponent of the bacterial cell wall, for example a component that isimportant in antigen pattern recognition and activating an optimaltargeted response. In comparing the 3 formulations containing phenol asa preservative (i.e., heat killed, phenol killed and radiation killed),irradiated K. pneumoniae antigenic composition led to the greatestrecruitment of acute inflammatory monocytes, DCs, NK cells, and T cellsto the lungs, followed by phenol killed K. pneumoniae antigeniccomposition, with heat killed K. pneumoniae antigenic compositionresulting in the least cellular recruitment.

As illustrated in FIGS. 4A to 4B, mice were treated on day 0, 2, 4, and6 with K. pneumoniae antigenic composition inactivated by heat (HKWP) orwithout (HKnp) phenol preservative, inactivated with phenol with (PKWP)or without (PKnp) phenol preservative, or K. pneumoniae antigeniccomposition inactivated by irradiation with phenol preservative (IRWP).On day 7 the mice were sacrificed and the total numbers of (A)inflammatory monocytes (CD11b+ Gr-1+) and DCs (CD11c+ lab+) or (B) CD4 Tcells, CD8 T cells, and natural killer (NK) cells were determined byflow cytometry. Error bars represent the sd of values from 5 mice pergroup. *p-value<0.05 compared to mice treated with IRWP using aStudent's t-test.

Example 2: Site Specificity Studies

With a focus on investigating the M1/M2 phenotypes in the in vivo modeldescribed herein which is used in conjunction with the antigeniccompositions described herein, the following experiments were performed.Briefly, 5 mice per group were treated on day 0, 2, 4, and 6 with eitherPBS, E. coli colon antigenic compositions, or K. pneumoniae antigeniccompositions. On day 7 of the experiment, the mice were sacrificed and abronchoalveolar lavage was performed. Subsequently the lungs andproximal colon were removed and enzymatically digested. After digestion,the recovered cells were washed and stained with antibodies specific forI-A/I-E FITC (MHC class II; M5/114.15.2); anti-Gr-1 PE (RB6-8C5_;anti-CD11 b PerCP-Cy5 (M1/70); anti-CD11c APC (N418). All antibodieswere acquired from eBioscience (San Diego, Calif.). The lung cells werecounted to determine the total number of cells (the colon was notcounted because we did not remove equal amounts of colon betweensamples). After staining for 20 mins the cells were washed and analyzedby FACS. Each data point shown in corresponding FIG. 5 represents thefrequency of CD11b+ Gr-1+ cells in the live gate for one mouse. As shownin FIG. 5, treatment with E. coli antigenic compositions leads to anincreased frequency of inflammatory monocytes in the colon of treatedmice.

Further, and as shown in FIGS. 6A to 6B, when monocytes in the lungswere examined based on the experimental methods detailed herein, it wasfound that while both E. coli and K. pneumoniae antigenic compositionsincrease the frequency of monocytes in the lungs of mice, K. pneumoniaeantigenic compositions were more effective when counting for totalnumbers. Referring to FIGS. 6A to 6B, the left-most panel shows thefrequency of CD11b+ Gr-1+ (inflammatory monocyte) cells in the lungs;the right-most panel shows the total number of CD11b+ Gr-1+ cells in thelung.

Example 3: Microbial Prophylaxis in Lungs

Example 3a: Klebsiella pneumoniae-derived antigenic formulation (SSI)protects against S. pneumoniae challenge in the lungs.

This Example illustrates that a SSI preparation of whole killedKlebsiella pneumonia administered subcutaneously in mice inducedincreases in circulating monocytes and provides protection againstsubsequent bacterial challenge with Streptococcus pneumoniae introducedinto the nasopharynx. Streptococcus pneumoniae, or pneumococcus, is aGram-positive, alpha-hemolytic, aerobic member of the orderLactobacillales. In contrast, Klebsiella pneumoniae is a Gram-negative,non-motile, encapsulated, lactose-fermenting, facultative anaerobemember of the order Enterobacteriales. These organisms are classified indistinct taxanomic phyla.

C57BL/6 mice were pretreated by subcutaneously injection with placebo orthe Klebsiella pneumonia formulation every other day for three weeks,then challenged with 1×10⁹ CFU of S. pneumonia P1547 and monitored for 5days for signs of clinical infection. On day 5, mice were sacrificed(moribund mice were sacrificed before day 5) and evaluated for bacterialload and immune parameters.

As illustrated in FIG. 7A, mice treated with the Klebsiella pneumoniaformulation were protected from S. pneumonia infection (5/5) whereasplacebo-treated mice were only partially protected (3/5). Furthermore,TNF and MCP-1 gene expression in lungs of resistant mice (both SSI- andplacebo-treated) were higher than placebo-treated mice that succumbed toinfection. FIG. 7B illustrates reduced bacterial counts in nasal cavity,lungs, and spleen with the SSI treatment.

Example 3b: Klebsiella pneumoniae-derived antigenic formulation (SSI)protects against P. aeruginosa or S. pneumoniae challenge in the lungs.

Pseudomonas aeruginosa (PA14, 7.8×10⁸ CFU/mouse) was instilled intolungs of 8-10 week old C57Bl/6 mice pretreated with 30 μL PBS or an SSIformulated from whole killed Klebsiella pneumoniae every other day for 3weeks, with survival as shown in FIG. 9A. Of the 6 surviving mice at day5, 3/6 of the PBS treated group had bacteria in the lungs and only 2/10SSI treated mice had bacteria in the lungs, as shown in FIG. 9B.

Streptococcus pneumoniae (P1542, 4.1×10⁶ CFU/mouse) was instilled intolungs of 8-10wo C57Bl/6 mice pretreated with 30 μL PBS or an SSIformulated from whole killed Klebsiella pneumoniae every other day for 3weeks, with survival as shown in FIG. 10.

This Example illustrates that a SSI preparation of whole killedKlebsiella pneumonia administered subcutaneously in mice inducesprophylactic antimicrobial activity against infection in the lungs withPseudomonas aeruginosa or S. pneumoniae. FIGS. 9A to 9B illustratesignificantly enhanced survival in a Pseudomonas aeruginosa challenge,and FIG. 10 illustrates enhanced survival in a S. pneumoniae challenge.These data illustrate prophylaxis mediated by an antigenic compositionof one lung pathogen, effective against two heterologous lung pathogens.

Example 3c: Comparison of Klebsiella pneumoniae-derived antigenicformulation (SSI) and E. coli-derived antigenic formulation (SSI) inprotecting against S. pneumoniae or P. aeruginosa challenge in thelungs.

In this Example, Klebsiella pneumoniae SSI (QBKPN) demonstratedstatistically superior efficacy in prophylaxis compared to the E. coliSSI (QBECO), in protecting against S. pneumoniae or P. aeruginosachallenge in the lungs.

For the model treatment of P. aeruginosa, mice were treated with theindicated SSI (0.03 ml/injection) for 14 days, then challenged withPseudomonas aeruginosa (PA14) by intranasal instillation of 6.0×10⁸ CFUof bacteria. Three days later, lungs were aseptically resected,homogenized, and assessed for bacterial load using Pseudomonas selectionagar plates. FIG. 15 shows the results (data are CFU/g of lung(mean+/−std dev), illustrating Klebsiella pneumoniae SSI (QBKPN)demonstrated statistically superior efficacy in prophylaxis compared tothe E. coli SSI (QBECO), in protecting against P. aeruginosa challengein the lungs.

For the model treatment of S. pneumoniae, mice were treated with theindicated SSI (0.03 ml/injection) for 14 days, then challenged withStreptococcus pneumoniae (PA14) by intranasal instillation of 5.0×10⁵CFU of bacteria. Three days later, lungs were aseptically resected,homogenized, and assessed for bacterial load using Pseudomonas selectionagar plates. FIG. 16 shows the results (data are CFU/g of lung(mean+/−std dev), illustrating Klebsiella pneumoniae SSI (QBKPN)demonstrated statistically superior efficacy in prophylaxis compared tothe E. coli SSI (QBECO), in protecting against S. pneumoniae challengein the lungs.

Example 4: Antimicrobial Therapy in GIT

This example illustrates heterologous anti-microbial therapy in a mousemodel of inflammatory bowel disease that uses a specific species (NRG857) of Adherent Invasive E. coli (AIEC) to induce a chronic IBD-likeinfection in the colon in the 129e strain of mice that have a reducedability to clear AIEC.

The 129e mice were infected with NRG 857 and treated with either (1)placebo, (2) a whole killed cell formulation of E. coli (SSI-1 in FIGS.8A to 8B), or (3) a whole killed cell formulation of S. enterica, whichis an exogenous pathogen which can cause gastrointestinal infection inmice (referred to as SSI-2 in FIG. 8). The results illustrate that thewhole killed formulation of E. coli was effective in reducing counts ofNRG 857 in colon tissue (300 times less than placebo treated mice—notethat the graph is a logarithmic scale) and feces of 129e mice. Inaddition, the S. enterica formulations also exhibited someanti-microbial activity against the heterologous AIEC infection.

In accordance with one aspect of the invention, macrophage defect ordeficiency, leading to a reduced ability to clear bacterial infectionand necrotic debris, may be the underlying trigger for some pathologiesassociated with microbial infections. This has for example beenpostulated to be the case in Crohn's disease. Aspects of the inventionaccordingly involve the induction of organ specific macrophagerecruitment and activation, resulting in clearance of bacterialinfection.

Example 5: Antimicrobial Prophylaxis in Peritoneal Cavity

In a first aspect, this example illustrates that a whole killed E. coliSSI formulation is effective in protecting against a S. enterica. Asillustrated in FIGS. 11A to 11B, SSI treated mice had significantlyfewer bacteria in the spleen than saline treated mice (p<0.0001), andSSI treated mice had less peak weight loss than saline treated mice(p<0.02).

In an alternative aspect, this Example illustrates that two alternativewhole killed E. coli SSI formulations (QBECO and QBECP), made fromdifferent strains of E. coli, are effective in a mouse model inprotecting against a S. enterica challenge, with levels of prophylaxisthat compare favourably to prior administration of an antigenic S.enterica formulation (QBSEN). In this Example, mice were pre-treated byskin injection with the alternative formulations, QBECO, QBECP (SSI fromurologic E. coli), or QBSEN (each given every other day for 3 weeks) andthen infected with S. enterica. Survival was improved for alltreatments, with QBECP providing comparable benefit to QBSEN, asillustrated in FIG. 12A. Mice were sacrificed, and colony counts in thespleen were quantitated and found to be lower in all the treated groups,as illustrated in FIG. 12B. Similarly, weight loss was found to be lowerin all the treated groups, as illustrated in FIG. 12C.

In a further illustration of targeted and optimized intraperitoneal (IP)prophylaxis, mice were treated with alternative SSIs, QBKPN and QBECO,or vehicle; then challenged with Salmonella enterica (typhimurium) by IPinstillation of 1.0×10⁶ CFU of bacteria. Three days later, spleens wereaseptically resected, homogenized, and assessed for bacterial load onHektoen enteric agar plates. FIG. 19 illustrates that both QBECO andQBKPN are protective, as measured by bacterial load in the spleen, andcounts in mice treated with QBECO are significantly lower than micetreated with QBKPN (data are CFU/ml (mean+/−std dev).

Example 6: Antimicrobial Prophylaxis in Skin

This example illustrates targeted heterologous anti-microbial therapy ina mouse model of skin infection, illustrating the improved efficacy ofantigenic formulations derived from microbial pathogens of the targettissue. Mice were treated with selected antigenic formulations of theinvention (0.03 ml/injection) for 14 days, then challenged withPseudomonas aeruginosa (PA14) by intradermal injection of 6.5×10⁵ CFU.Three days later, skin was aseptically resected, homogenized, andassessed for bacterial load using Pseudomonas selection agar plates.FIG. 13 illustrates that QBSAU (S. aureus-derived SSI) protects againstP. aeruginosa challenge in skin, with significantly reduced bacterialcounts following QBSAU, compared to QBKPN (which is not a skin pathogenin the murine model). A repeat iteration of the foregoing procedurereplicated the results, as shown in FIG. 14 (n=8 mice/group).

Example 7: Antimicrobial Prophylaxis in Geriatric Model Aged Mice

This example illustrates targeted heterologous anti-microbial therapy ina geriatric mouse model of lung infection, showing that Klebsiellapneumoniae SSI (QBKPN) protects against S. pneumoniae challenge in lungsof aged mice. FIGS. 17A to 17B show that the survival benefit for oldmice, FIG. 17B, was more pronounced than for young mice, FIG. 17A. FIG.18 illustrates corroborative data, showing that QBKPN reduces weightloss in aged mice, following challenge with S. pneumoniae, and thisbenefit is greater for aged mice compared to young mice.

Example 8: Antiviral Prophylaxis in Lungs

In this Example, mice wherein injected with QBKPN or QBECO SSI, orvehicle for 14 days, in accordance with the protocol outlined in otherExamples. Mice were then challenged intranasally with murine herpesvirus 68 (MHV68), a virus that is genetically modified to drive aluciferase reporter. MHV68 is a model viral pathogen for studying lunginfection. Three days after challenge, mice were injected with 300micrograms of luciferin; 10 min later, mice were sacrificed, lungsresected, and imaged with a Xenogen IVIS imager. Luminescence in thelungs is an indicator of active viral infection and replication.

Pretreatment of mice with QBKPN SSI dramatically diminished theluminescence signal in lungs of mice, providing evidence of antiviralprophylaxis in lungs using targeted antigenic bacterial formulations.The same dramatic result was not observed for QBECO.

OTHER EMBODIMENTS

Although various embodiments of the invention are disclosed herein, manyadaptations and modifications may be made within the scope of theinvention in accordance with the common general knowledge of thoseskilled in this art. Such modifications include the substitution ofknown equivalents for any aspect of the invention in order to achievethe same result in substantially the same way. In some embodiments, theinvention excludes steps that involve medical or surgical treatment.Numeric ranges are inclusive of the numbers defining the range. In thespecification, the word “comprising” is used as an open-ended term,substantially equivalent to the phrase “including, but not limited to”,and the word “comprises” has a corresponding meaning. Citation ofreferences herein shall not be construed as an admission that suchreferences are prior art to the present invention. All publications areincorporated herein by reference as if each individual publication werespecifically and individually indicated to be incorporated by referenceherein and as though fully set forth herein. The invention includes allembodiments and variations substantially as hereinbefore described andwith reference to the examples and drawings.

1-69. (canceled)
 70. A method of modulating an immune system in avertebrate host for the prophylactic treatment of infection by a firstmicrobial pathogen in a target tissue, the method comprising:administering to the vertebrate host an effective amount of an antigenicformulation comprising antigenic determinants specific for a secondheterologous microbial pathogen, wherein: the second heterologousmicrobial pathogen is an Escherichia coli; the antigenic determinantsare selected from the group consisting of whole cells or cell wallextracts; and, the first microbial pathogen is: a gastrointestinal tractpathogen, and the target tissue is the gastrointestinal tract; or, apathogen of a peritoneal cavity organ, and the target tissue is theperitoneal cavity; wherein the administering comprises administeringsuccessive doses of the formulation at a dosage interval of at least onehour, so that two or more doses are administered over a period from 2days to 1 month, over a dosage duration of at least one week.
 71. Themethod of claim 70, wherein the first microbial pathogen is thegastrointestinal tract pathogen, and the target tissue is thegastrointestinal tract.
 72. The method of claim 70, wherein the firstmicrobial pathogen is the pathogen of the peritoneal cavity organ, andthe target tissue is the peritoneal cavity.
 73. The method of claim 70,wherein the host is a human patient.
 74. The method of claim 70, whereinthe antigenic determinants comprise whole cells.
 75. The method of claim73, wherein the antigenic determinants comprise whole cells.
 76. Themethod of claim 74, wherein the whole cells are killed.
 77. The methodof claim 75, wherein the whole cells are killed.
 78. The method of claim74, wherein the whole cells are attenuated.
 79. The method of claim 75,wherein the whole cells are attenuated.
 80. The method of claim 70,wherein the antigenic determinants comprise cell wall extracts.
 81. Themethod of claim 73, wherein the antigenic determinants comprise cellwall extracts.